Source: UNIVERSITY OF ARKANSAS submitted to
UNDERSTANDING FUNDAMENTAL MECHANISMS OF FEED EFFICIENCY IN POULTRY
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
1006354
Grant No.
(N/A)
Project No.
ARK02479
Proposal No.
(N/A)
Multistate No.
(N/A)
Program Code
(N/A)
Project Start Date
Apr 27, 2015
Project End Date
Apr 26, 2020
Grant Year
(N/A)
Project Director
Bottje, WA.
Recipient Organization
UNIVERSITY OF ARKANSAS
(N/A)
FAYETTEVILLE,AR 72703
Performing Department
Poultry Sciences
Non Technical Summary
This population in the world will exert an expected demand for high quality animal protein of approximately 75% more than is being currently produced today. Because there will only be an increase of 10% of arable land used for grain production that will be used for animal production, there is a huge demand for increased productive efficiency in animal agriculture. One component of improved animal production efficiency is the inherent genetic capability of animals to convert plant products into high quality animal protein otherwise known as feed efficiency. In addition, the major cost (60 to 70%) of raising an animal to market weight is due to the cost of feed. Consequently, understanding fundamental mechanisms of feed efficiency is of vital importance to develop tools that can help commercial animal agriculture breeding continue to improve feed efficiency in their genetic stock. A final issue that the world is facing that can have serious negative impact on feed efficiency are increasing temperatures attributed to climate change. In the last decade, the world has experienced some the highest global temperatures on record. Heat stress, just as any stress that an animal may experience, can have negative consequences on health and productivity. Therefore, studies outlined in this proposal will investigate effects that heat stress can have on fundamental cellular mechanisms including feed efficiency.To investigate fundamental mechanisms of feed efficiency, we will use a two pronged approach. One approach will be to study targeted genes, proteins, or metabolic pathways that can be affected by heat stress, or chemical stress that can induce oxidative stress. Cells have natural defenses to combat stress that include antioxidant protection or pathways that can help correct damage to cell components that can be caused by outside effects (e.g. heat stress) or effects that can be mediated by the cells themselves (e.g. oxidative stress). There are reactive chemicals inside cells that can be produced through normal metabolism. We will use different cell lines to investigate specific mechanisms to determine how these metabolic pathways might be altered by heat or oxidative stress. Unlike animals, it is much easier to change expression of targeted genes or proteins using specific chemical inhibitors or activators, or by manipulation of the amount of specific gene or proteins. By nature, these cellular based studies offer a high degree of control of targeted pathways that can not be accomplished in whole animal studies.Ultimately however, to have true biological meaning in animal agriculture, mechanisms that may be revealed in cell culture studies such as those described above, must be tested, analyzed, and delineated in animals. Therefore, we will conduct research in animals exposed to control conditions (e.g. ideal temperature conditions, called thermoneutral conditions) as well as under high ambient temperature conditions (i.e. heat stress conditions. During these studies, various tissues such as muscle, liver, intestines, brain and adipose tissue will be obtained after humane euthanasia of animals. These tissues then be analyzed for specific molecules (e.g. proteins, fats) and genes to understand the effects of heat stress on specific metabolic pathways. Animal studies in this project will be conducted using commercial broilers (meat chickens) but will also utilize unique lines of quail that have been specifically selected or greater or lesser susceptibility to stress. By imposing an external heat stress on top of this inherent difference in stress response in these birds, we anticipate that we will reveal new insight into how animals respond to environmental stress. Another animal model that will be utilized in these studies are quail that are being selected for high feed efficiency or low feed efficiency. Exposure of these animals to heat stress can reveal how temperature stress can adversely affect growth and fundamental mechanisms of feed efficiency. The ultimate goal will be try to help producers continue to grow poultry efficiently even under high environmental temperatures
Animal Health Component
0%
Research Effort Categories
Basic
70%
Applied
20%
Developmental
10%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3053220102025%
3053220104025%
3053299102025%
3053299104025%
Knowledge Area
305 - Animal Physiological Processes;

Subject Of Investigation
3220 - Meat-type chicken, live animal; 3299 - Poultry, general/other;

Field Of Science
1040 - Molecular biology; 1020 - Physiology;
Goals / Objectives
1) To determine fundamental cellular and molecular mechanisms associated with the phenotypic expression of feed efficiency in poultry.2) To determine how cellular mechanisms of feed efficiency are altered in response to exogenous and endogenous oxidative stress.
Project Methods
Studies associated with this project include live animal investigation followed by tissue sampling and analysis (Approach 1) as well as experiments conducted in cell culture (Approach 2) in which conditions can be very tightly controlled and/or manipulated more readily than in whole animal studies.Approach 1: Animal studies that are planned will use commercial broilers raised under thermoneutral or heat stress conditions. Broilers will be provided standard control diets or diets supplemented with different levels of various antioxidants (e.g. tocopheral, vitamin C, plant extracts known to contain high levels of antioxidants). In addition, studies will be conducted using a unique line of Japanese Quail that have been selected for susceptibility (S) or resistance (R) stress based on levels of circulating corticosterone levels after a period of restraint. In addition, research will be conducted using a line of quail that is being divergently selected for high feed efficiency or low feed efficiency. As quail reach sexual maturity very quickly (6 to 7 wks), it is anticipated that analysis of tissues obtained from these animals how changes in targeted cellular mechanisms/metabolic pathways may change with continued selection pressure for feed efficiency. Comparisons of feed efficiency in these lines can also be compared to the quail line being selected for feed efficiency as well.Approach 2: The second general approach will be to conduct heat and oxidative stress studies in specific cell lines (e.g. avian-derived muscle, liver, fibroblasts). In these studies, cells could be treated with chemical agents (e.g. antioxidants) or given a period of heat stress (e.g. exposure to high temperatures for varying time periods) after which cells will be harvested and analyzed for gene and protein expression analysis. Fundamental cellular mechanisms such as bioenergetics, post-translational modification, cell death, etc., can also be elucidated by using methodology to increase or decrease expression of targeted genes or proteins. This methodology could include chemical inhibitors or activators, gene transfection (to increase gene expression), or small inhibitory RNA (to inactivate or inhibit) targeted molecules or pathways.Evaluation: A measure of success of the project will be various outputs that include presentations at scientific meetings and publications of research in peer reviewed journals.

Progress 04/27/15 to 04/26/20

Outputs
Target Audience:The target audience for this project are poultry industry and allied poultry industry scientists as well as students and faculty scientists in academia. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Two students (Kentu Lassiter and Alissa Piekarski) received training and successfully completed their respective PhD programs. Another student (Devin Cook) completed an MS program. There were also opportunities provided to undergraduate students who worked on projects in the lab. How have the results been disseminated to communities of interest?The results have been disseminated to scientists in the field through the peer reviewed publications provided in this report. In addition, numerous presentations were made at scientific meetings. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? The body of work contained in these studies represents the development of a very comprehensive understanding of the cellular basis of feed efficiency based on the global gene and protein expression studies that had been conducted. These studies were conducted primarily in muscle tissue of pedigree line of male broilers. This line would have a great influence on the genetic makeup of commercial broilers. The studies culminated in several conference presentations made at meetings of the Poultry Science Association as well as the American Society of Animal Science.

Publications

  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Lassiter, K., E. Greene, O. B. Faulkner, B. M. Hargis, W. Bottje, and S. Dridi. 2015. Orexin system is expressed in avian muscle cells and regulates mitochondrial-related genes. Am. J. Physiol. 308(3):R173-87. doi: 10.1152/ajpregu.00394.2014. Epub 2014 Dec 10. Piekarski, A., N. B. Anthony, W. Bottje, S. Dridi. 2015. Crosstalk between Autophagy and obesity: potential use of avian model. Advances in Food Technology http://dx.doi.org/10.17140/AFTNSOJ-1-106 Greene, E., K. Lassiter, W. Bottje, S. Dridi. 2015. Orexin regulates mitochondrial dynamics in avian muscle tissue. Receptors and Clinical Investigation. e632. doi: 10.14800/rci.632 Nguyen, P., E. Greene, P. Ishola, G. Huff, A. Donoghue, W. Bottje, S. Dridi. 2015. Chronic mild cold conditioning modulates expression of hypothalamic neuropeptide and intermediary metabolic related genes and improves growth performance in young chicks. PLoS One 11/2015 10(11):e0142319.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Kong, B-W., K. Lassiter, A. Piekarski, S. Dridi, A-Reverter-Gomez, N. Hudson, and W. Bottje. 2016. Proteomics of breast muscle tissue associated with the phenotypic expression of feed efficiency within a pedigree male broiler line. I. Highlight on mitochondria. PLoS 11(5): e0155679. doi:10.1371/journal.pone.0155679
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Reverter, A., R. Okimoto, R. Sapp, W.G. Bottje, R. Hawken, and N.J. Hudson. 2016. Chicken muscle mitochondrial content appears coordinately regulated and is associated with performance phenotypes. Biology Open 2016: doi:10.1242/bio.02272
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: 121. Bottje, W.G., K. Lassiter, S. Dridi, N. Hudson, B-W. Kong. 2017. Enhanced expression of proteins involved in energy production and transfer in breast muscle of pedigree male broilers exhibiting high feed efficiency. Poult. Sci. 96:2454-2458. Doi: 10.3382/ps/pew453.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W.G., B-W. Kong, A. Reverter, A. J. Waardenberg, K. Lassiter, and N. J. Hudson. 2017a. Progesterone signaling in broiler skeletal muscle is associated with divergent feed efficiency. BMC Systems Biology (2017) 11:29 DOI 10.1186/s12918-017-0396-2.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: 125. Bottje, W.G., K. Lassiter, A. Piekarski-Welsher, S. Dridi, A. Gomez-Reverter, N.J. Hudson, and B-W. Kong. 2017b. Proteogenomics reveals enriched ribosome assembly and protein translation in Pectoralis major of high feed efficiency pedigree broiler males. Frontiers in Physiology. 8 (article 306):1-11. Doi: 10.3389/fphys.2017.00306.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Kong, B-W., N. Hudson, D. Seo, S. Lee, B. Khatri, K. Lassiter, D. Cook, A. Piekarski, S. Dridi, N. Anthony, and W. Bottje. 2017. RNA sequencing for global gene expression associated with muscle growth in a single male modern broiler line compared to a foundational Barred Plymouth Rock chicken line. BMC Genomics 18:82 DOI: 10.1186/s12864/s12864-016-3471.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W. G., B. Khatri, S. A. Shouse, D. Seo, B. Mallmann, S. K. Orlowski, J. Pan, S. Kong, C. M. Owens, N. B. Anthony, J. K. Kim, and B. C. Kong. Identification and differential abundance of mitochondrial genome encoding small RNAs (mitosRNA) in breast muscles of modern broilers and unselected chicken breed. Frontiers in Physiology 8:816.doi: 103389/fphys.2017.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Hudson, N. J., W. G. Bottje, R. J. Hawken, B. Kong, R. Okimoto and A. Reverter. 2017. Mitochondrial metabolism a key driver of energy utilization and product quality? Animal Prod. Sci. 57:22042215
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Rodriques, D. R., K. M. Wilson, M. Trombetta, W. N. Briggs, A. F. Duff, K. M. Chasser, W. G. Bottje, and Lisa Bielke. 2020. A proteomic view of the cross-talk between early intestinal microbiota and poultry immune system. Front. Physiology-Avian Physiology. Front. Physiol. https/doi:10.3389/fphys.2020.00020
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Piekarski-Welsher, A., E. Greene, K. Lassiter, B. C. Kong, S. Dridi, and W. Bottje. 2018. Enrichment of autophagy and proteosome pathways in breast muscle of feed efficient pedigree male broilers. Frontiers in Physiology doi: 10.3389/fphys.2018.01342.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: 132. Lassiter, K., S. Dridi, E. Greene, B. Kong, and W.G. Bottje. 2018. Identification of mitochondrial hormone receptors in avian muscle cells. Poult. Sci. 97:2926-2933.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Bottje, W. G. 2019. Oxidative metabolism and efficiency: The delicate balancing act of mitochondria. Poult. Sci. 98:4223-4230.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Lassiter, K. B. C. Kong, A. Piekarski-Welsher, S. Dridi, and W. G. Bottje. 2019. Gene expression essential for myostatin signaling and skeletal muscle development is associated with divergent feed efficiency in pedigree male broilers. Front. Physiol. 26 February 2019. https://doi.org/10.3389/fphys.2019.00126
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Bottje, W. G. Oxidative stress and efficiency: the tightrope act of mitochondria in health and disease (Board Invited Review). 2019. J. Anim. Sci. 97:3169-3179 doi: 10.1093/jas/skz219


Progress 10/01/18 to 09/30/19

Outputs
Target Audience:The target audience is the academic science community for fundamental research as well as the poultry industry - primarily commercial breeding companies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?One student, Kentu Lassiter, completed his PhD and is now serving in a post-doctoral role on a major 5 year USDA-NIFA project in sustainability in agriculture. How have the results been disseminated to communities of interest?The research has been reported at scientific meetings and published in peer reviewed journals. What do you plan to do during the next reporting period to accomplish the goals?We will continue to investigate global expression analysis associated with feed efficiency as well as wooden breast muscle myopathy.

Impacts
What was accomplished under these goals? Review papers by Bottje (2019) tied together a large number of studies focused on the cellular basis of feed efficiency in broiilers. A faculty member from another University who had reviewed this paper indicated, 'I REALLY enjoyed your paper! I use several other works of yours in my Energetics class!'. To me, this is the greatest compliment that could be given to my research in feed efficiency. Targeted gene expression analysis in breast muscle of the high feed efficiency (FE)pedigree male (PedM) broilerphenotype reported by Lassiter et al. (2019) revealed that muscle development may be fostered in the high FE PedM phenotype by down-regulation several components of the myostatin signaling pathway genes combined with upregulation of genes that enhance muscle formation and growth. There was also evidence of genetic architecture that would foster muscle protein synthesis in the high FE phenotype. A clear indication of differences in insulin signaling between high and low FE phenotypes was not apparent in this study. These findings indicate that a gene expression architecture is present in breast muscle of PedM broilers exhibiting high FE that would support enhanced muscle development-differentiation as well as protein synthesis compared to PedM broilers exhibiting low FE. The study by Mallmann et al. (2019,aimed to assess the fat infiltration in bone marrow and its repercussion on broiler chicken affected by Wooden Breast (WB) myopathy. Ten Cobb 500 live birds with extreme rigidity of the Pectoralis major (PM) muscle were selected as WB affected chickens by physical examination of the muscle at 49 days of age, whereas ten chickens healthy with no physical signs of hardness in the breast muscle were considered to be unaffected. Macroscopic lesions in affected chickens included areas of firm and inflamed muscle with pale appearance, hemorrhaging, and viscous exudate on the surface. Bone marrow and sections of the PM muscle were collected and analyzed for light microscopy. Additionally, transmission electron microscopy was conducted in affected or unaffected muscle. Chickens affected with WB showed significant reductions (P < 0.05) in femur diameter, calcium, and phosphorous percentage but increased breast weight, compression force and filet thickness when compared with non-affected chickens. Interestingly, bone marrow from WB chicken had subjectively, more abundant infiltration of adipose tissue, when compared with non-affected chickens. Histology of the Pectoralis major of birds with WB showed abundant infiltration of adipose tissue, muscle fibers degeneration with necrosis and infiltration of heterophils and mononuclear cells, connective tissue proliferation, and vasculitis. Ultrastructural changes of WB muscle revealed lack definition of bands in muscle tissue, or any normal ultrastructural anatomy such as myofibrils. The endomysium components were necrotic, and in some areas, the endomysium was notable only as a string of necrotic tissue between degraded myofibrils. Tissue changes in WB chickens appeared hypertrophied, with large areas of necrosis and myofiber without structural identity with degraded mitochondria adjacent to the disrupted muscle tissue. As far as we know, this is the first study that describes a subjective increase in adipose tissue in the bone marrow of chickens affected with WB when compared with non-affected chickens, and reduced bone mineralization.

Publications

  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Bottje, W. G. 2019. Oxidative metabolism and efficiency: The delicate balancing act of mitochondria. Poult. Sci. 98:4223-4230.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Lassiter, K. B. C. Kong, A. Piekarski-Welsher, S. Dridi, and W. G. Bottje. 2019. Gene expression essential for myostatin signaling and skeletal muscle development is associated with divergent feed efficiency in pedigree male broilers. Front. Physiol. 26 February 2019. https://doi.org/10.3389/fphys.2019.00126
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Bottje, W. G. Oxidative stress and efficiency: the tightrope act of mitochondria in health and disease (Board Invited Review). 2019. J. Anim. Sci. 97:3169-3179 doi: 10.1093/jas/skz219
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Mallmann, B. A., E. M. Martin, K. S. Kim, N. L. Calderon-Apodaca, M. F. Baxter, J. D. Latorre, X. Hernandez-Velasco, L.Paasch-Martinez, C. M. Owens, W. G. Bottje, E. S. Greene, G. Tellez. Evaluation of bone marrow adipose tissue and bone mineralization on broiler chickens affected by wooden breast myopathy. Front. Physiol. Avian Phys. May 2019 10, article 674 doi:10.3389/fphys:2019.000674
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Kozaczek, M. W. Bottje, E. Greene, K. Lassiter, B. Kong, S. Dridi, S. Korourian, and R. Hakkak. Comparison of liver gene expression by RNAseq and PCR analysis after 8 weeks of feeding soy protein isolate- or casein-based diets in an obese liver steatosis rat model. Food & Function. DOI:10.1039/c9fo01387
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Bottje, W., V. Kuttappan, C. M. Owens, K. Lassiter, and B. Kong. 2019. Comparison of upstream regulator analysis of wooden breast proteomics in commercial broilers with breast muscle proteomics in Pedigree Male Broilers exhibiting high feed efficiency. Poult. Sci. 98(E): 171
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lassiter, K., V. Kuttappan, C. M. Owens, B. C. Kong, S. Dridi, and W. Bottje. 2019. Wooden breast myopathy upstream regulator analysis of proteomic data. Poult. Sci. 98(E):171.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Rodrigues, D. R., K. Wilson, W. Briggs, A. Duff, K. Chasser, J. Bielke, W. Bottje, and L. Bielke. 2019. A proteomic view of cross-talk between intestinal microbiota and host immune system. Poult. Sci. 98(E): 99
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Briggs, W., D. R. Rodriques, K. Wilson, A. Duff, K. Chasser, J. Bielke, W. Bottje, and L. Bielke. 2019 Impact of intestinal pioneer colonizers on inflammation in broiler chicks at ten days of age. 2019 Poult. Sci. 98(E): 179


Progress 10/01/17 to 09/30/18

Outputs
Target Audience:The target audience would primarily be the scientific community and personnel in the poultry industry. Changes/Problems:Comparison of proteomic data in feed efficiency to that in broilers with muscle myopathy is an addition to the project. The reason for this change is that there is considerable interest in the commercial poultry industry to try to understand and eliminate this muscle myopathies as they are costly to the commercial integratorsin terms of down grades at the poultry processing plants. What opportunities for training and professional development has the project provided?A minoirty student has completed a PhD in this program. How have the results been disseminated to communities of interest?The information has been presented at scientific conferences and has been published in peer reviewed journals. What do you plan to do during the next reporting period to accomplish the goals?We will continue to explore information that is contained in global gene and protein expression datasets. We are also planning to compare a feed efficiency proteomic dataset to proteomic data in a type of muscle myopathy. This investifation will help shed light on how selection of feed efficency may or may not contribute to muscle myopathy in commercial broilers.

Impacts
What was accomplished under these goals? We have added considerable knowledge towards understanding fundamental mechanisms of feed efficiency in breast muscle in a unique feed efficiency animal; pedigree male broilers.

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: 1) Bottje, W. G. 2018. Oxidative metabolism and efficiency: The delicate balancing act of mitochondria. Poult. Sci. (Oct. https://doi.org/10.3382/ps/pey405 ). 2)Piekarski-Welsher, A., E. Greene, K. Lassiter, B. C. Kong, S. Dridi, and W. 1) 1) Bottje. W. 2018. Enrichment of autophagy and proteosome pathways in breast muscle of feed efficient pedigree male broilers. Frontiers in Physiology October 2018 doi: 10.3389/fphys.2018.01342 3) Piekarski, A., G. Nagarajan, P. Inole, J. Flees, E. S. Greene, W. J. Kuenzel, T. Onkubo, H. Maier, W. G. Bottje, M. A. Cline, S. Dridi. 2018. AMP-Activated protein kinase mediates the effect of leptin on avian autophagy in a tissue-specific manner. Frontiers in Physiology  Avian Physiology 9:1-17. Doi:10.3389/ fphys.2018.00541.


Progress 10/01/16 to 09/30/17

Outputs
Target Audience:The target audience for research conducted in this project would be the scientific community in animal agriculture, government and academic scientists, and personnel in commercial breeding companies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A PhD student (Alissa Piekarski-Welsher) was able to be successfully funded USDApost-doctoral fellowship as a direct result of some of the research conducted on this project as a PhD student. She took a position as a research scientist in a company in allied industry. Kentu Lassiter (PhD student) is projected to complete his PhD in May 2018. Kentu has been serving in the capacity as program associate (technician) intimately involved in most of the research presented above. How have the results been disseminated to communities of interest?The results have been published in peer reviewed journals, a book chapter, and several invited presentations at conferences. What do you plan to do during the next reporting period to accomplish the goals?There are 4 manuscripts in progress that continue to build more background in understanding cellular mechanisms of feed efficiency.

Impacts
What was accomplished under these goals? Increased mitoproteome expression in high feed efficiency muscle. In two separate global expression studies (transcriptomic and proteomic) we reported that the numbers of genes and proteins associated with the mitoproteome is elevated in muscle of animals with high feed efficiency. This complements previous research conducted in this area indicating an enhanced mitochondrial content. Progesterone Signaling Using Regulatory Impact Factor (RIF) analysis it was determined that progesterone signaling is playing a role in the phenotypic expression of feed efficiency. We believe that this signaling mechanism is taking place in an autocrine mechanism within muscle of broilers. From this finding we conducted research that indicates that avian mitochondria express hormone receptors for progesterone, estrogen, corticosterone, thyroid hormone and insulin - all of which could have profound effects on bioenergetics in cells. Enhanced energy production and transfer in high FE muscle. Enhanced 'machinery' for energy production and energy transfer was revealed in high FE muscle. In cells with fluctuating energy demand (e.g. skeletal muscle), a transfer system of proteins across the inner and outer mitochondrial membranes links mitochondrial oxidative phosphorylation to cytosolic phosphorylated creatine (PCr) that serves as a phosphate reservoir for rapid repletion of cytosolic ATP. Crucial proteins of this energy transfer system include several creatine kinase (CK) isoforms found in the cytosol and mitochondria.From these findings, we hypothesize that enhanced expression of the energy production and transfer system in breast muscle of the high FE pedigree broiler male could be fundamentally important in the phenotypic expression of feed efficiency.

Publications

  • Type: Book Chapters Status: Published Year Published: 2017 Citation: Bottje, W. G. and B-W. Kong. 2017. The cellular basis of feed efficiency in poultry muscle: mitochondria and nucleic acid metabolism. In: Achieving sustainable production of poultry meat. Ed. Todd Applegate. Burleigh Dodds Series in Agricultural Science (No. 14) (p. 51-69).
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W.G., K. Lassiter, S. Dridi, N. Hudson, B-W. Kong. 2017. Enhanced expression of proteins involved in energy production and transfer in breast muscle of pedigree male broilers exhibiting high feed efficiency. Poult. Sci. 96:2454-2458. Doi: 10.3382/ps/pew453.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Kuttappan, V.; W. Bottje, R. Ramanathan, S. Hartson, C. Coon, B-W. Kong, C. Owens, M. Vazquez-Ano, and B. Hargis. 2017. Proteomic analysis reveals changes in carbohydrate and protein metabolism associated with broiler breast myopathy. Poult. Sci. 96: 2992-2999.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W.G., B-W.Kong, A. Reverter, A. J. Waardenberg, K. Lassiter, and N. J. Hudson. 2017. Progesterone signaling in broiler skeletal muscle is associated with divergent feed efficiency. BMC Systems Biology 11:29 DOI 10.1186/s12918-017-0396-2.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W.G., K. Lassiter, A. Piekarski-Welsher, S. Dridi, A. Gomez-Reverter, N.J. Hudson, and B-W. Kon. 2017. Proteogenomics reveals enriched ribosome assembly and protein translation in Pectoralis major of high feed efficiency pedigree broiler males. Frontiers in Physiology. 8 (article 306):1-11. Doi: 10.3389/fphys.2017.00306.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: 6) Kong, B-W., N. Hudson, D. Seo, S. Lee, B. Khatri, K. Lassiter, D. Cook, A. Piekarski, S. Dridi, N. Antony, and W. Bottje. 2017. RNA sequencing for global gene expression associated with muscle growth in a single male modern broiler line compared to a foundational Barred Plymouth Rock chicken line. BMC Genomics 18:82 DOI: 10.1186/s12864/s12864-016-3471.y
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Bottje, W. G., B. Khatri, S. A. Shouse, D. Seo, B. Mallmann, S. K. Orlowski, J. Pan, S. Kong, C. M. Owens, N. B. Anthony, J. K. Kim, and B. C. Kong. 2017. Identification and differential abundance of mitochondrial genome encoding small RNAs (mitosRNA) in breast muscles of modern broilers and unselected chicken breed. Front. Physiol. (published Oct. 20, 2017) 8:1- 9. doi 10:3389/fphys.2017.00816.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Hudson, N. J., W. G. Bottje, R. J. Hawken, B. Kong, R. Okimoto and A. Reverter. 2017. Mitochondrial metabolism a key driver of energy utilization and product quality? Animal Prod. Sci. 57:22042215
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Bottje, W. 2017. Efficiency and oxidative metabolism: The difficult balance. Oxidative Stress Symposium. Poultry Science Assoc. July 20, 2017.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Bottje, W. 2017. Global expression studies associated with muscle growth in Pedigree Male Broilers and Barred Plymouth Rock. European Egg and Meat Conference. Edinburgh UK Sept 4-6, 2017.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Proteomics of feed efficiency: Mitochondria and more. Arkansas Bioscience Annual Meeting. Oct. 25, 2017. Fayetteville, AR
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Bottje, W. 2017. Proteogenomics reveals enriched ribosome assembly and protein translation in Pectoralis major of high feed efficiency pedigree broiler males. Nutrition Conference. Oct. 26. 2017, Fayetteville, AR.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2017 Citation: Bottje, W. 2017. Revealing mechanisms of muscle myopathy with pathway analysis. Graduate Student Workshop. Univ. of Georgia. Oct. 7, 2017
  • Type: Other Status: Other Year Published: 2017 Citation: Bottje, W. 2017. Proteogenomics of Feed Efficiency. Iowa State University  Seminar program. April 14, 2017. Des Moines, IA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Khatri, B. S. Shouse, D. Seo, N. Hudson, W. Bottje, B. Kong. 2017. MicroRNA profiling associated with muscle growth in modern broiler compared to unselected Barred Plymouth Rock chickens. Poult. Sci
  • Type: Conference Papers and Presentations Status: Other Year Published: 2017 Citation: Lassiter, K., S. Dridi, E. Greene, B. Kong, and W. Bottje. 2017. The presence of hormone receptors in the mitochondria of avian muscle cells. Poult. Sci.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Bottje, W., B-W. Kong, and N. J. Hudson. 2017. Proteogenomics provides evidence of enhanced ribosome assembly, and protein translation in muscle of high vs low feed efficiency pedigree broiler males. 2017. Plant and Animal Genome Meeting


Progress 10/01/15 to 09/30/16

Outputs
Target Audience:The target audience for this outcomes of this project would be the scientific community in animal agriculture as well as government scientists and personnel in commercial animal breeding companies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The research conducted on this project directy contributed to Dr. Alissa Piekarski-Welscher completing her PhD program and lead to her successful funding of a USDA Post-Doctoral Fellowship that she received in March of 2016. Devin Cook completed his master's thesis which was a part of this overall program. Kentu Lassiter is a PhD candidate whose project is being funded in part by this program. How have the results been disseminated to communities of interest?The results of our studies have been presented at scientific meetings and is being published in different scientific journals as well as a book chapter that will be published in March 2017. What do you plan to do during the next reporting period to accomplish the goals?We will continue to investigate fundamental mechanisms of feed efficiency by exploiting the unique proteogenomic datasets that we have generated in these studies.

Impacts
What was accomplished under these goals? INTRODUCTION We have conducted global gene expression (RNAseq) and global protein expression (shotgun proteomics) on muscle tissueobtained from animals exhibiting high and low feed efficiency (FE) phenotypes. Whereas the RNAseq study was part of theoriginal proposal objective, the proteomics investigation was an 'add on' to the project. From these studies, and another globalgene expression study conducted on the same set of tissue samples, we now have a unique capability to exploit theproteogenomic data to develop an understanding of the cellular basis of feed efficiency in a way that no other research group(to our knowledge) has been able to do. SPECIFIC RESULTS 1) Increased expression of mitoproteome. In both Bottje et al. (2016a) and Kong et al. (2016)., we determined that thenumbers of genes and proteins associated with the mitoproteome is biased towards the high FE group, suggesting a slightlyelevated mitochondrial content in high FE. This complements previous research conducted in this area indicating anenhanced mitochondrial content. 2) Role of progesterone in feed efficiency. In Bottje et al. (2017), we applied a differential network method (RegulatoryImpact Factors) whose aim is to use patterns of differential co-expression to detect regulatory molecules transcriptionallyrewired between the groups. This analysis clearly points to alterations in progesterone signaling (via the receptor PGR) as themajor driver, and we show the progesterone receptor localizes to the mitochondria in a quail muscle cell line. Progesterone issometimes used in the cattle industry in exogenous hormone mixes that lead to a ~20% increase in FE. Because theprogesterone receptor can localize to avian mitochondria, our data continue to point to muscle mitochondrial metabolism asan important component of the phenotypic expression of variation in broiler FE. 3)Enhanced energy production and transfer in high FE muscle. In Bottje et al. (2016b), evidence was presented indicatingthe presence of enhanced 'machinery' for energy production and energy transfer of in high FE muscle tissues. In cells withfluctuating energy demand (e.g. skeletal muscle), a transfer system of proteins across the inner and outer mitochondrialmembranes links mitochondrial oxidative phosphorylation to cytosolic phosphorylated creatine (PCr) that serves as aphosphate reservoir for rapid repletion of cytosolic ATP. Crucial proteins of this energy transfer system include severalcreatine kinase (CK) isoforms found in the cytosol and mitochondria. In a recent proteomic study (Bottje et al., 2016a), severalcomponents of this system were upregulated in high FE compared to low FE breast muscle; notably adenine nucleotidetranslocase (ANT), voltage dependent activated channel (VDAC), the brain isoform of creatine kinase (CK-B), and severalproteins of the electron transport chain. Reexamination of the original proteomic dataset revealed that the expression of twomitochondrial CK isoforms (CKMT1A and CKMT2) had been detected but were not recognized by the bioinformatics programused by Kong et al. (2016a). The CKMT1A isoform was up-regulated (7.1 fold, P = 0.05) in the high FE phenotype but therewas no difference in CKMT2 expression (1.1 fold, P = 0.59). From these findings, we hypothesize that enhanced expressionof the energy production and transfer system in breast muscle of the high FE pedigree broiler male could be fundamentallyimportant in the phenotypic expression of feed efficiency. 4)Enhanced capability for RNA and DNA synthesis and metabolism in high FE. Evidence from global gene and proteinexpression studies (proteogenomics) was presented by Bottje and Kong (2016) indicating that in breast muscle obtained frompedigree male broilers exhibiting high or low feed efficiency phenotypes, there is evidence of enhanced mitochondrialfunction, enhanced mitochondrial energy (ATP) production, transfer, and cellular energy reserve (phospho-creatine),enhanced for synthesis and metabolism of purine and pyrimidine nucleotides that could lead to enhanced ability to construct RNA and DNA in muscle. 5) Identification of mitochondrial hormone receptors in avian muscle. Based on results of Bottje et al. (2016) and Kong et al. (2016) indicating that progesterone would be playing an important role in the phenotypic expression of feed efficiency, studies were conducted that identified the presence of hormone receptors for progesterone, glucocorticoid and thyroid hormone on mitochondria (Lassiter et al., 2016). This study was conducted to determine the presence of receptors for progesterone (PR), glucocorticoid (GR), thyroid (TR), and insulin (IR) in nuclear, cytosolic and mitochondrial cellular fractions in an avian muscle cell line (quail muscle 7, QM7) and in breast muscle of quail and broilers. Visualization of receptor location in QM7 cells was accomplished by immunofluorescence. QM7 cells were stained with Mito Tracker Deep Red CMX, fixed in methanol, immune stained with anti-PR, -GR, -TR, and -IR primary antibodies overnight at 4°C, and visualized with Alexa Fluor 488-conjugated secondary antibody. After staining the nucleus with DAPI, images were obtained by immuno-fluorescence microscopy. The results indicate that all hormone receptors were detected in the nucleus and cytosol in QM7 cells. Merged images clearly revealed PR, GR, and TR in mitochondria (indicated by yellow to orange color) of QM7 cells with evidence of mitochondrial IR as well. Differential centrifugation was used to confirm the presence of hormone receptors on mitochondria in QM7 cells and in breast muscle obtained from quail and broilers. In QM7 cells, antibody binding of the mitochondrial fraction was observed for PR, IR, and TR, but not GR. The presence of all 4 receptors were detected in the mitochondrial fraction in breast musclefrom quail and broilers. To our knowledge, this is the first report of hormone receptors (PR, TR, GR and IR) on mitochondriain avian muscle cells. We hypothesize that these receptors could play important direct roles in regulating avian mitochondrialfunction. 6)Comparison of modern (efficient) broiler line to heritage breed. By investigating global gene expression in a highly selected pedigree broiler line and a foundational breed (Barred Plymouth Rock), the results provide insight into cellular mechanisms that regulate muscle growth, fiber composition and feed efficiency.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Kong, B-W., K. Lassiter, A. Piekarski, S. Dridi, A-Reverter-Gomez, N. Hudson, and W. Bottje. 2016. Proteomics of breast muscle tissue associated with the phenotypic expression of feed efficiency within a pedigree male broiler line. I. Highlight on mitochondria. PLoS 11(5): e0155679. doi:10.1371/journal.pone.0155679
  • Type: Book Chapters Status: Awaiting Publication Year Published: 2017 Citation: Bottje, W. G. and B-W. Kong. Cellular basis of feed efficiency in muscle: Focus on mitochondria and nucleic acid metabolism. Chapter 6 In: Achieving sustainable production of poultry meat. Burleigh Dodds Publishing Ltd, Cambridge CB223HJUK
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2016 Citation: Reverter, A., R. Okimoto, R. Sapp, W. Bottje, R. Hawken, and N. Hudson. 2016. Chicken muscle mitochondrial content appears coordinately regulated and associated with performance phenotypes. Biol. Open: doi:10.1242/bio.02272
  • Type: Journal Articles Status: Accepted Year Published: 2017 Citation: Bottje, W., B-W. Kong, A. Reverter, A. J. Waarendenburg, K. Lassiter, and N. J. Hudson. 2017. Progesterone signalling in broiler skeletal muscle is associated with divergent feed efficiency. BMC Systems Biol. (in press).
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Sharifabadi, H. R., Naserian, A. A., Valizadeh, R. Nassiry, M. R., Bottje, W. G., Renden, R. R. 2016. Growth performance, feed digestibility, body composition, and feeding behavior of high and low-residual feed intake fat-tailed lambs under moderate feed restriction. J. Anim. Sci. 94:3382-3388.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Baum, J. I., T.A. Washington, S.A. Shouse, W. Bottje, S. Dridi, G. Davis, D. Smith. 2016. Leucine supplementation at the onset of high-fat feeding does not prevent or improve glycemic regulation in male Sprague-Dawley rats. J. Biochem. Physiol. DOI: 10.1007/s13105-016-0516-2 (Aug. 20, 2016).
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Abousaad, S., K. Lassiter, A. Piekarski, P. Chary, K. Striplin, K. Christensen, L.R. Bielke, B. M. Hargis, S. Dridi, and W. Bottje. 2017. Effects of in ovo feeding of dextrin-iodinated casein in broilers. I. Hatch weights and early growth performance. Poult. Sci. (Http://dx.doi.org/10.3382/ps/pew439 ).
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Abousaad, S., K. Lassiter, A. Piekarski, P. Chary, K. Striplin, K. Christensen, L.R. Bielke, B. M. Hargis, S. Dridi, and W. Bottje. 2017. Effects of in ovo feeding of dextrin-iodinated casein in broilers. II. Hatch window and growth performance. Poult. Sci. (http://dx.doi.org/10.3383/ps/pew438 ).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Cook, D., B. Kong, K. Lassiter, E. Greene, A. Piekarski, S. Dridi, and W. Bottje. 2016. Expression of Nuclear Factor [Erythroid-derived 2]-like 2 (NFE2L2) and Kelch like-ECH protein 1 (KEAP1) in breast muscle of broiler breeder males with high or low feed efficiency. Poult. Sci. 95:(E-Suppl.1): 253.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Bottje, W.G., K. Lassiter, S. Dridi, N. Hudson, B-W. Kong. 2017. Enhanced expression of proteins involved in energy production and transfer in breast muscle of 4 pedigree male broilers exhibiting high feed efficiency. Poultry Sci. (in press).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: K. Lassiter, S. Dridi, E. Greene, N. Hudson, B. Kong, L. Berghman, and W. Bottje. 2016. Evidence of hormone receptors in mitochondria of QM7 cells and breast muscle of quail and broilers. Poult. Sci. 95 (E-Suppl. 1): 47
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: 4. Kong, B-W., D. Seo, N. Hudson, B. Khatri, and W. Bottje. 2016. Bioinformatic integration of two independent studies on gene expression of breast muscles in modern broiler pedigree chickens compared with unselected heritage chickens. Poult. Sci. 95:(E-Suppl. 1):168.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Bottje, W. K. Lassiter, S. Dridi, N. Hudson, and B-W. Kong. 2016. Hormonal and mitochondrial signatures in muscle: Possible implications for mitochondrial function and feed efficiency. Poult. Sci. 95(E-Suppl 1):75.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: 9. Rajaei-Sharifabadi, H., E. Greene, K. Lassiter, A. Piekarski, D. Cook, K. Blankenship, P. Nguyen, A. Decker, L. Gramlich, Y. Thaxton, Y. Yiang, L. Ellestad, T. Porter, W. Bottje, and S. Dridi. 2016. Noni (Morinda citrifolin) modulates the hypothalamic expression of feeding-related neuropeptides and heat shock proteins in broilers exposed to acute heat stress. Poult. Sci. 95(E-Suppl. 1):149.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Decker, A., E. Greene, B-W. Kong, and S. Dridi. 2016. Expression of neuropeptide Y system in avian liver and its role in hepatic lipogenesis. Poult. Sci. 95(E-Suppl. 1):134.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: 11. Lassiter, K. B.W. Kong, S. Dridi, and W. Bottje. 2016. Genes involved in muscle development, muscle differentiation, and insulin signaling may play a role in the phenotypic expression of feed efficiency in a single genetic line of male broilers. Poult. Sci. 95(E-Suppl. 1):197.


Progress 04/27/15 to 09/30/15

Outputs
Target Audience:The target audience for this project will be the scientific community involved in production animal agriculture and for geneticists working for commercial animal breeding companies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?One graduate student is working on the project towards an MS degree with an anticipated completion date of August, 2016. A second graduate student completed a PhD and is now working as a post-doctoral fellowship that was funded by a grant she wrote to USDA-NIFA. How have the results been disseminated to communities of interest?The information has been disseminated at national meetings and has also been published in peer reviewed journals. What do you plan to do during the next reporting period to accomplish the goals?We are planning to continue working in this area to understand cellular mechanisms associated with feed efficiency. We have some very exciting preliminary results that suggests that there are hormone receptors on avian muscle cell mitochondria. While there is evidence of the presence of receptors on mitochondria in mammalian cells, to our knowledge, this has not been reported in avian cells.

Impacts
What was accomplished under these goals? The publications that were cited revealed that the hormone orexin, previously considered to be only secreted by the hypothalamus to regulate feed intake, was actually shown to be produced by muscle cells and has an impact on cell bioenergetics. As such, this opens up a new mechanism of cellular bioenergetics that could have important ramifications on the phenotypic expression of feed efficiency. Research was also conducted that demonstrated a role of autophagy ('self eating') in the phenotypic expression of feed efficiency. Autophagy is a mechanism used by cells to eliminate damaged components of the cell (e.g. molecules, organelles). Surprisingly, it appears that autophagy may be more active in broilers that are more feed efficient, suggesting that the ability to correct/fix damaged components allows the cells to operate more efficiently. This process might be best described as an analogy to regular maintenance of a car. If one takes a car in for regular maintenance, there is an expenses that comes from changing oil, rotating tires etc. If someone chooses not to have regular maintance, no expense occurs in the short term initially. However, major expenses will be occured if the car is not maintained. Similarly, increased autophagy may serve as a regular maintenance expense to fix damaged components of the cell but it enables the cell to function optimately. Therefore, increased autophagy could help make animals more efficient by allowing cells to function optimally.

Publications

  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Journal Articles: Lassiter, K., E. Greene, O. B. Faulkner, B. M. Hargis, W. Bottje, and S. Dridi. 2015. Orexin system is expressed in avian muscle cells and regulates mitochondrial-related genes. Am. J. Physiol. 308: R173R187. Greene, E., K. Lassiter, W. Bottje, S. Dridi. 2015. Orexin regulates mitochondrial dynamics in avian muscle tissue. Receptors and Clinical Investigation. e632. doi: 10.14800/rci.632. Dridi, S., N. Anthony B-W. Kong, and W. Bottje. 2015. Feed Efficiency: A key production trait and a global challenge. Adv.Food Tech. Nutr. Sci. http://dx.doi.org/10.17140/AFTNSOJ-1-e004 Piekarski, A., N. B. Anthony, W. Bottje, S. Dridi. 2015. Crosstalk between Autophagy and obesity: potential use of avian model. Advances in Food Technology http://dx.doi.org/10.17140/AFTNSOJ-1-106 Abstracts: Piekarski, A., W. Bottje, B-W. Kong, S. Dridi. 2015. Role of Autophagy and its effect on feed efficiency in commercial poultry. Autophagy Meeting. Keystone, CO. June 2015. Blankenship, K., A. Gilley, E. Greene, N. Anthony, B-W. Kong, N. Hudson, W. Bottje, S. Dridi. Expression profile of feeding-related hypothalamic neuropeptides in quail lines selected for high or low feed efficiency. Poult. Sci.94(1): 15. Lassiter, K., B.W. Kong, S. Dridi, W. Bottje. 2015. Investigating differential gene and protein expression to understand the cellular basis of feed efficiency in commercial broilers. Poult. Sci. 94(1):15. Kong, B-W Kong, B. Khatri, K. Lassiter, D. Cook. A. Piekarski, N. Anthony, S. Dridi, N. Hudson, W. Bottje. 2015. Gene expression of breast muscle tissue associated with the phenotypic expression of feed efficiency in a single male modern broiler line compared to progenitor Barred Plymouth Rock broiler line. Poult. Sci. 94(1): 164-165.