THE FUNCTIONAL ROLE OF SREBP1C AND S14 IN REGULATION OF MAMMARY LIPID SYNTHESIS BY RUMEN-DERIVED BIOACTIVE FATTY ACIDS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: AFRI COMPETITIVE GRANT
• Reporting Frequency: Annual
• Accession No.: 0221493
• Grant No.: 2010-65206-20723
• Cumulative Award Amt.: $349,476.00
• Proposal No.: 2009-05586
• Project Start Date: Apr 15, 2010
• Project End Date: Apr 14, 2015
• Grant Year: 2010
• Program Code: [92220]- Animal Growth and Nutrient Utilization

Project Director
Harvatine, K.

Recipient Organization
PENNSYLVANIA STATE UNIVERSITY
208 MUELLER LABORATORY
UNIVERSITY PARK,PA 16802

Performing Department
Animal Science

Non Technical Summary
Milk is an important source of nutrients and fat is the most variable milk component. Nutrition has a major impact on milk fat yield in the dairy cow, the most striking example being milk fat depression (MFD), which is commonly observed in cows fed certain diets. Our previous work has established that unique fatty acids produced as intermediates in the rumen during biohydrogenation of dietary polyunsaturated fatty acids regulate milk fat synthesis. Our goal is to develop an understanding of the regulation of milk fat synthesis by these naturally occurring bioactive fatty acids. The work proposed uses an integrated approach to understand the molecular regulation of milk fat synthesis by investigating the functional mechanisms of previously identified regulatory pathways. Specifically, we will first characterize the regulation of a key enzyme of lipid synthesis, acetyl-CoA carboxylase (ACC), test the functional role of two previously identified transcription factors (SREBP1 and S14), and further investigate the role of endoplasmic reticulum stress. This investigation will identify the functionally important signals regulating fat synthesis in the mammary gland. The long-term goal of this project is to increase the efficiency of milk fat synthesis by improving the understanding of the biological mechanisms underlying synthesis of milk fat. The knowledge gained will allow development of management strategies to improve productivity and animal well-being, optimize producer return and sustainability, and enhance quality of dairy products for consumers.

Animal Health Component

20%

Research Effort Categories

Basic

80%

Applied

20%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
302	3410	1010	75%
302	3840	1010	25%

Knowledge Area
302 - Nutrient Utilization in Animals;

Subject Of Investigation
3840 - Laboratory animals; 3410 - Dairy cattle, live animal;

Field Of Science
1010 - Nutrition and metabolism;

Keywords

milk fat

milk fat depression

biohydrogenation intermediate

conjugated linoleic acid

lipogenesis

srebp1

spot 14

er stress

lipid synthesis

acetyl-coa carboxylase

Goals / Objectives
The work proposed specifically focuses on the molecular regulation of milk fat depression by investigating the functionality and mechanisms of identified regulatory pathways. Specific aims include: 1. Investigate regulation of acetyl-CoA carboxylase and fatty acid synthase during milk fat depression. 2. Establish the mechanism of SREBP-1c and S14 in the regulation of milk fat synthesis by fatty acids derived from rumen biohydrogenation. 3. Determine if endoplasmic reticulum stress reduces milk fat synthesis through modification of ACC activity or abundance. The long-term goal of this project is to increase the efficiency of milk fat synthesis by improving the understanding of the biological mechanisms underlying lactation and metabolism. The propose work will provide strong insight into the regulation of milk fat synthesis in the dairy cow and the functional importance of candidate regulatory mechanisms.

Project Methods
An integrated approach using a combination of in vitro and in vivo experiments will be used to investigate transcriptional and post-translational responses during milk fat depression and the functional importance of candidate regulatory systems. We will first characterize the isoform specific expression and post-translational regulation of acetyl-CoA carboxylase (ACC) during milk fat depression. The bioactive fatty acid responsive region of the ACC promoter will be characterized using a reporter gene approach and site-specific mutagenesis. The functional importance of regulation of ACC and fatty acid synthase (FASN) during milk fat depression will then be tested using chemical inhibitors and over-expression of ACC in mammary epithelial cell culture. A central role and physiological relevance of thyroid hormone responsive spot 14 (S14) and sterol response element binding protein 1 (SREBP1) in this regulation by fat synthesis during milk fat depression will be first be tested in the S14 null and mammary SREBP1 activation null (floxed SCAP + beta-lactoglobulin driven CRE) mice. Secondly, we will investigate if over-expression of S14 and SREBP1 rescues biohydrogenation intermediate inhibition of fat synthesis in mammary epithelial cell culture. Lastly, we will investigate if bioactive fatty acid induced endoplasmic reticulum (ER) stress reduces fat synthesis through modification of ACC activity or abundance using chemical inhibitors and activators of ER stress signaling and over-expression of tribbles homolog 3 (TRB3) and activating transcription factor 4 (ATF4).

Progress 04/15/10 to 04/14/15

Outputs
Target Audience:The target audience reached was scientists through presentations and publications and industry nutritionist, consultants and upper level managers through papers and nutrition conference presentations. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?During the past year the project has supported research of PhD graduate students, one post-doctoral scholar, and a technician. Additionally, undergraduate students were involved as technical assistants. How have the results been disseminated to communities of interest?Original research has been disseminated through peer reviewed journal articles, review articles, and abstracts at a national conferences. Additionally, the PD was invited to speak on milk fat synthesis at an international conference in Beijing China and industry conferences in the United States. 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 work specifically focused on the molecular regulation of milk fat depression by investigating the functionality and mechanisms of identified regulatory pathways. Specific aims included: 1. Investigate regulation of acetyl-CoA carboxylase and fatty acid synthase during milk fat depression. 2. Establish the mechanism of SREBP-1c and S14 in the regulation of milk fat synthesis by fatty acids derived from rumen biohydrogenation. 3. Determine if endoplasmic reticulum stress reduces milk fat synthesis through modification of ACC activity or abundance. The long-term goal of this project is to increase the efficiency of milk fat synthesis by improving the understanding of the biological mechanisms underlying lactation and metabolism. The proposed work provided strong insight into the regulation of milk fat synthesis in the dairy cow and the functional importance of candidate regulatory mechanisms. Maximal milk fat yield remains an important variable to farm profitability and milk fat depression continues to be a real-world problem on dairy farms. This project has focused on the role of trans-10, cis-12 conjugated linoleic acid (CLA) in induction of milk fat depression. We have determined that activation of Sterol Response Element Binding Protein 1 (SREBP1) is essential to inhibition of milk fat synthesis using a transgenic mouse model with a mammary specific deletion of a gene essential to SREBP1 activation. We also demonstrated that Thyroid Hormone Responsive Spot 14 (S14) is essential to determining the extent of response to CLA as the S14 null mouse is hyper-responsive, but not hypersensitive to CLA. We have used promoter specific quantitative Real-Time PCR assays for splice variants of acetyl-CoA carboxylase to determine the specific regulation of this key enzyme of fat synthesis during milk fat depression. We have observed that although some pathways of ER stress are activated during milk fat depression, chemical activation of endoplasmic reticulum stress does not reduce fat synthesis in mammary epithelial cells. These experiments have provided a mechanistic test of important regulatory pathways in the mammary gland and their response to bioactive fatty acids. The project provided insight into managing and reducing milk fat depression directly applicableon farms and basic insight into the regulatory mechanism of the condition. The project also significantly contributed to our long term goal to develop a complete understanding of the regulation of milk fat synthesis and apply this knowledge in the development of management strategies to improve productivity and animal well-being, optimize economic return and sustainability of producers, and enhance quality of dairy products for consumers. Specifically, the project was successful in demonstrating the functional importance of two regulatory systems and moved this knowledge beyond previous correlations. Establishing the functional role of SREBP1 and S14 in milk fat synthesis is key to allowing investment in future experiments that target these pathways to reduce milk fat depression and stimulate milk fat yield. Additionally, the project demonstrated the ability to integrate experimental observations in the cow and transgenic mice to allow investigation not feasible directly in the cow. The findings in the project provide a platform for the development of methods to enhance milk fat yield and improve the FA profile of milk fat. For example, SREBP1, SREBP1 regulatory proteins, and S14 have been used as candidate genes for identification of single nucleotide polymorphisms (SNPs) that may explain genetic differences in milk fat yield and FA composition of bovine fat. The project provided insight into managing and reducing milk fat depression directly applicableon farms and basic insight into the regulatory mechanism of the condition. The project also significantly contributed to our long term goal to develop a complete understanding of the regulation of milk fat synthesis and apply this knowledge in the development of management strategies to improve productivity and animal well-being, optimize economic return and sustainability of producers, and enhance quality of dairy products for consumers. Specifically, the project was successful in demonstrating the functional importance of two regulatory systems and moved this knowledge beyond previous correlations. Establishing the functional role of SREBP1 and S14 in milk fat synthesis is key to allowing investment in future experiments that target these pathways to reduce milk fat depression and stimulate milk fat yield. Additionally, the project demonstrated the ability to integrate experimental observations in the cow and transgenic mice to allow investigation not feasible directly in the cow. The findings in the project provide a platform for the development of methods to enhance milk fat yield and improve the FA profile of milk fat. For example, SREBP1, SREBP1 regulatory proteins, and S14 have been used as candidate genes for identification of single nucleotide polymorphisms (SNPs) that may explain genetic differences in milk fat yield and FA composition of bovine fat.

Publications

• Type: Journal Articles Status: Published Year Published: 2015 Citation: Rico, D.E., S.H Preston, J.M. Risser, and K.J. Harvatine. Key rumen microbial populations are rapidly changed during induction of and recovery from diet-induced milk fat depression in dairy cows. Bri. J. Nutr.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Ma, L., K.L. Cook, D.E. Bauman, and K.J. Harvatine. 2014. Inhibition of milk fat synthesis during CLA- and diet-induced milk fat depression occurs equally across the day in Holstein cows. J. Dairy Sci. 98:1851-1855.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Niu, M., Y. Ying, P.A. Bartell, K.J. Harvatine. 2014. The effects of feeding time on the circadian pattern of feed intake, milk production, and plasma hormones and metabolites in dairy cows. J. Dairy Sci. 97:7764-76.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Harvatine, K.J., M.M. Robblee, S.R. Thorn, Y.R. Boisclair, and D.E. Bauman. 2014. Trans-10, cis-12 conjugated linoleic acid dose-dependently inhibits milk fat synthesis without disruption of lactation in C57BL/6J mice. J. Nutr. 144:1928-1934.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Rico, D.E., Y. Ying, and K.J. Harvatine. 2014. Comparison of enriched palmitic acid and calcium salts of palm fatty acid fat supplements on milk production and metabolic profiles of dairy cows. J. Dairy Sci. 97:5637-5644.
• Type: Journal Articles Status: Published Year Published: 2013 Citation: Rico, D.E. and K.J. Harvatine. 2013. Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration. J. Dairy Sci 96:6621-6630.
• Type: Journal Articles Status: Published Year Published: 2013 Citation: Hussein, M., K. H. Harvatine, W. M. P. B. Weerasinghe, L. A. Sinclair, and D. E. Bauman. 2013. Conjugated linoleic acid-induced milk fat depression in lactating ewes is accompanied by reduced expression of mammary genes involved in lipid synthesis. J. Dairy Sci. 96:3825-3834
• Type: Journal Articles Status: Published Year Published: 2011 Citation: Harvatine, K.J. and D.E. Bauman. 2011. Characterization of the acute lactational response to trans-10, cis-12 conjugated linoleic acid (CLA). J. Dairy Sci. 94:6047-6056.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Harvatine, K.J. 2015. Recent advances in nutrient gene interactions regulation milk fat synthesis. Beijing China. May 9-11.
• Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Harvatine, K.J. 2013. Fatty acid composition and fat levels in milk. Proceedings of the 3rd International Symposium on Dairy Cow Nutrition and Milk Quality. pg. 1-8. Beijing China. May 11-13.
• Type: Other Status: Published Year Published: 2013 Citation: Harvatine, K.J. 2013. What is old and new in dietary fat for dairy cows? Farmshine Magazine. May 3
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Harvatine, K.J. 2012. Causes of diet induced milk fat depression and strategies to recover. Proceedings of the Mid-South Ruminant Nutrition Conference. Grapevine, TX. April 25-26.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Harvatine, K.J. 2012. Circadian patterns of feed intake and milk component variability. Proceedings of the Tri-State Dairy Nutrition Conference. Fort Wayne, IN. pp. 34-54.
• Type: Other Status: Published Year Published: 2013 Citation: Oliveira, D.E., K.J. Harvatine, Y.R. Boisclair, and D.E. Bauman. 2013. PPARgamma agonists and antagonists fail to overcome trans-10, cis-12 conjugated linoleic acid (CLA) inhibition of lipogenesis and lipogenic gene expression in bovine mammary epithelial cell culture. J. Dairy Sci. 96(E-Suppl. 1):325.
• Type: Other Status: Published Year Published: 2012 Citation: Harvatine, K.J., M. Tanino, Y.R. Boisclair, and D.E. Bauman. 2012. Thyroid hormone responsive spot 14 null mice are acutely responsive to trans-10, cis-12 conjugated linoleic acid (CLA) in the mammary gland. J. Dairy Sci. 95(E-Suppl. 2):416.

Progress 04/15/13 to 04/14/14

Outputs
Target Audience: Nutrition and lactation researchers and dairy nutritionists. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? During the past year the project has supported research of two PhD graduate students, two post-doctoral scholar, and. Additionally, one undergraduate students hascompleted an independent projects and an additional 6 undergraduate students were involved as technical assistants. How have the results been disseminated to communities of interest? Original research has been disseminated through peer reviewed journal articles, review articles, and abstracts at a national conferences. Additionally, the PD was invited to present at an international nutrition techniques symposium in Brazil and industry conferences in the United States. What do you plan to do during the next reporting period to accomplish the goals? During the next year the remainder of the laboratory and data analysis will be conducted. Additionally, manuscripts will be completed for publication.

Impacts
What was accomplished under these goals? Maximal milk fat yield remains an important variable to farm profitability and milk fat depression continues to be a real-world problem on dairy farms. This project has focused on the role of trans-10, cis-12 conjugated linoleic acid (CLA) in induction of milk fat depression. We have determined that activation of Sterol Response Element Binding Protein 1 (SREBP1) is essential to inhibition of milk fat synthesis using a transgenic mouse model with a mammary specific delection of an gene essential to SREBP1 activation. We also demonstrated that Thyroid Hormone Responsive Spot 14 (S14) is essential to determining the extent of response to CLA as the S14 null mouse is hyper-responsive, but not hypersensitive to CLA. We have used promoter specific quantitative Real-Time PCR assays for splice variants of acetyl-CoA carboxylase to determine the specific regulation of this key enzyme of fat synthesis during milk fat depression. We have observed that although some pathways of ER stress are activated during milk fat depression, chemical activation of endoplasmic reticulum stress does not reduce fat synthesis in mammary epithelial cells. Current ongoing work is investigating the effect of inhibition of specific rate limiting enzymes during bioactive treatment of mammary epithelial cells. These experiments have provided a mechanistic test of important regulatory pathways in the mammary gland and their response to bioactive fatty acids.

Publications

• Type: Journal Articles Status: Published Year Published: 2013 Citation: Rico, D.E. and K.J. Harvatine. 2013. Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration. J. Dairy Sci 96:6621-30.
• Type: Journal Articles Status: Published Year Published: 2013 Citation: Hussein, M., K.H. Harvatine, Y.R. Boisclair, and D.E. Bauman. 2013. Supplementation with fish oil as a source of n-3 fatty acids does not downregulate mammary lipogenesis in lactating mice. J. Nutr. 1913-9.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Rico, D.E., E.R. Marshall, J. Choi, K.E. Kaylegian, C.D. Dechow, and K. J. Harvatine. 2014. Within milking variation in milk composition and fatty acid profile of Holstein dairy cows. J. Dairy Sci. 97:1-10.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Harvatine, K.J., Y.R. Boisclair, D.E. Bauman. 2014. LXR stimulates lipogenesis in bovine mammary epithelial cell culture but does not appear to be involved in diet-induced milk fat depression in cows. Physiological Reports. 27:e00266.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Rico, D.E., Y. Ying, A.R. Clarke, and K.J. Harvatine. 2014. The effect of rumen digesta inoculation on the time course of recovery from diet-induced milk fat depression in dairy cows. J. Dairy Sci. 97:3752-3760.
• Type: Journal Articles Status: Accepted Year Published: 2014 Citation: Rottman-Gredell, L.W., Y. Ying, K. Zhou, P.A. Bartell, and K.J. Harvatine. Effect of the timing of feed intake on the daily rhythm of milk synthesis in the cow. Physiol. Reports. ACCEPTED.
• Type: Journal Articles Status: Accepted Year Published: 2014 Citation: Rico, D.E., Y. Ying, and K.J. Harvatine. 2014. Comparison of enriched palmitic acid and calcium salts of palm fatty acid fat supplements on milk production and metabolic profiles of dairy cows. J. Dairy Sci. ACCEPTED.
• Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Harvatine, K.J. 2014. Advances in measuring ruminal biohydrogenation and flux of fatty acids to the duodenum. Proceedings of the V International Symposium Advances on Research Techniques for Ruminant Nutrition. Pirassununga, Brazil. April 10-11.
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Rico, D.E., S.H. Preston, and K.J. Harvatine. 2014. Time-course of select ruminal microbes during induction and recovery from diet-induced milk fat depression in dairy cows. Accepted to ADSA-ASAS JAM, Kansas City, MO, July 20-24.
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Sandri, E.C., E.M. Sandri, E. Ticiani, M. Urio, M. Camera, A.P. Povaluk, K.J. Harvatine, D.E. de Oliveira. 2014. The peroxisome proliferator-activated receptor gamma (PPAR?) agonist thiazolidinedione (TZD) does not overcome trans-10, cis-12 conjugated linoleic acid (CLA) inhibition of milk fat synthesis in lactating dairy ewes. Accepted to ADSA-ASAS JAM, Kansas City, MO, July 20-24.
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Oliveira, D.E., D.E. Bauman, and K.J. Harvatine. 2014. Conjugated linoleic acid (CLA) trans-10, cis-12 decreases ACC-? gene expression in lactating mammary gland by decreasing specific transcripts from different promoters. Accepted to ADSA-ASAS JAM, Kansas City, MO, July 20-24.
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Harvatine, K.J. 2014. Insights provided by nutrigenomics into the effect of diet on metabolism and milk production. ADSA/ASAS JAM Triennial Lactation Symposium. Kansas City, MO. July 20-24, 2014.
• Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Harvatine, K.J. 2014. Milk Fat Recovery Interventions. Cargill Dairy Supplier Technology Exchange. Tannersville, PA. Jan. 7-8.
• Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Harvatine, K.J. 2013. Production and Use of Volatile Fatty Acids (VFAs) in Dairy Cattle. Penn State Dairy Nutrition Workshop. Grantville, PA. Nov. 12-13.
• Type: Other Status: Other Year Published: 2013 Citation: Harvatine, K.J. 2013. Recovering From Milk Fat Depression. Virtus Fatty Acid Forum. Webinar. July 18th 2013.
• Type: Journal Articles Status: Accepted Year Published: 2014 Citation: Jenkins, T. C. and K.J. Harvatine. 2014. Lipid feeding and milk fat depression. Veterinary Clinics of North America: Food Animal Practice. ACCEPTED.

Progress 04/15/12 to 04/14/13

Outputs
Target Audience: Nutrition and lactation researchers and dairy nutritionists. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? During the past year the project has supported research of two PhD graduate students, a post-doctoral scholar, and a visiting research scholar who where mentored by the PD. Additionally, two undergraduate students have completed independent projects and an additional 6 undergraduate students were involved as technical assistants. How have the results been disseminated to communities of interest? Four original research abstracts were presented at a national conference during 2012 and two abstracts are accepted for presentation during the summer of 2013. Additionally, the PD was invited to present at an international dairy symposium in Beijing China and an industry conferences in the United States. What do you plan to do during the next reporting period to accomplish the goals? During the next year the remainder of the laboratory and data analysis will be conducted. Additionally, manuscripts will be completed for publication.

Impacts
What was accomplished under these goals? Maximal milk fat yield remains an important variable to farm profitability and milk fat depression continues to be a real-world problem on dairy farms. This project has focused on the role of trans-10, cis-12 conjugated linoleic acid (CLA) in induction of milk fat depression. First, we determined that activation of Sterol Response Element Binding Protein 1 (SREBP1) is essential to inhibition of milk fat synthesis using a transgenic mouse model with a mammary specific delection of an gene essential to SREBP1 activation. We also demonstrated that Thyroid Hormone Responsive Spot 14 (S14) is essential to determining the extent of response to CLA as the S14 null mouse is hyper-responsive, but not hypersensitive to CLA. We have used promoter specific quantitative Real-Time PCR assays for splice variants of acetyl-CoA carboxylase to determine the specific regulation of this key enzyme of fat synthesis during milk fat depression. Lastly, we have observed that although some pathways of ER stress are activated during milk fat depression, chemical activation of endoplasmic reticulum stress does not reduce fat synthesis in mammary epithelial cells. These experiments have provided a mechanistic test of important regulatory pathways in the mammary gland and their response to bioactive fatty acids.

Publications

• Type: Conference Papers and Presentations Status: Accepted Year Published: 2013 Citation: D.E. Oliveira, K.J. Harvatine, Y.R. Boisclair, and D.E. Bauman. 2013. PPARgamma agonists and antagonists fail to overcome trans-10, cis-12 conjugated linoleic acid (CLA) inhibition of lipogenesis and lipogenic gene expression in bovine mammary epithelial cell culture. Accepted to ADSA/ASAS Joint Annual Meeting, Indianapolis, IN.
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2013 Citation: Ma, L., Y. Ying, A. R. Clarke, P. Bartell, and K. J. Harvatine. 2013. Characterization of mammary circadian rhythms of wild-type C57BL/6J mice and the role of Thyroid Hormone Responsive Spot 14 (S14) in circadian regulation of milk fat synthesis. Accepted to ADSA/ASAS Joint Annual Meeting, Indianapolis, IN.
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Rico, D.R. and K.J. Harvatine. 2012. Rescuing milk fat depression: Tackling the most important risk factors first. Penn State Dairy Nutrition Workshop. Grantville, PA. November 13-14.
• Type: Journal Articles Status: Under Review Year Published: 2013 Citation: Rico, D.E. and K.J. Harvatine. 2013. Induction of and recovery from milk fat depression occurs progressively in dairy cows switched between diets that differ in fiber and oil concentration. J. Dairy Sci. Submitted.
• Type: Journal Articles Status: Published Year Published: 2013 Citation: Conjugated linoleic acid-induced milk fat depression in lactating ewes is accompanied by reduced expression of mammary genes involved in lipid synthesis
• Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Harvatine, K.J. 2013. Fatty acid composition and fat level in milk. Proceedings of the 3rd International Symposium on Dairy Cow Nutrition and Milk Quality. May 11-13. Beijing, China. Pg. 1-8

Progress 04/15/11 to 04/14/12

Outputs
OUTPUTS: During the second year of the grant we published one peer reviewed paper, published four abstracts, presented at five regional conferences (Tri-State Dairy Nutrition Conference, Mid-South Ruminant Nutrition Conference, Mid-Atlantic Bovine Practitioners Conference, Penn State Dairy Nutrition Workshop, and Bucknell Nutrition Conference). Additionally, four abstracts have been accepted to national meetings for 2012. Investigation of the functional role of two transcription factors in induction of diet-induced milk fat depression has been completed. Investigation of the regulation of acetyl-CoA carboxylase during milk fat depression is ongoing. PARTICIPANTS: J. Moeller (PhD canidate) was a visiting scholar. Y. Ying and A. Clarke are research technicians. D. Rico is a third year PhD student. K. Cook and E. Marshall are undergraduates who conducted independent research projects. Other undergraduates were involved at multiple levels. TARGET AUDIENCES: Nutrition and lactation researchers and dairy nutritionists. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Milk is an economically and nutritionally important component of milk, but is very responsive to diet. We have determined the functional role of two candidate regulatory systems, SREBP1 and S14, in diet inhibition of milk fat synthesis. Our data supports an essential role for SREBP1 while S14 regulates the extent of milk fat depression. Additionally we have determined that a third candidate regulatory system (ER stress) appears to not be directly involved in diet induced milk fat depression. Understanding the functional role of regulatory systems in the dietary regulation of milk fat synthesis is essential to development of targeted selection and management systems to increase milk fat. Improved management of milk fat will reduce the environmental impact on dairy production and improve producer sustainability.

Publications

• Harvatine, K. J. and D. E. Bauman. 2011. Characterization of the acute lactational response to trans-10, cis-12 conjugated linoleic acid (CLA). J. Dairy Sci. 94:6047-56.
• Harvatine, K. J. 2012. Circadian Patterns of Feed Intake and Milk Composition Variability. Proceedings. Tri-State Dairy Nutrition Conference. Fort Wayne, Indiana. April, 2012. p. 43-55.
• Harvatine, K. J. 2012. Causes of Diet Induced Milk Fat Depression and Strategies To Recover. Proceedings Mid-South Ruminant Nutrition Conference. Dallas, TX. April, 2012. 8 pages.
• Hussein, M., K. J. Harvatine, W. M. P. B. Weerasinghe, L. A. Sinclair, D. E. Bauman. 2011. Conjugated linoleic acid-induced milk fat depression in lactating ewes is accompanied by reduced expression of genes involved in mammary lipid synthesis. J Dairy Sci. 94(E-Suppl. 1):M195.
• Rottman, L. W., Y. Ying, and K. J. Harvatine. 2011. Effect of timing of feed intake on circadian pattern of milk synthesis. J Dairy Sci. 94(E-Suppl. 1):830.
• Rico, D. E. and K. J. Harvatine. 2011. Effect of a high palmitic acid fat supplement on ruminal fermentation and milk production in high- and low-producing dairy cows. J Dairy Sci. 94(E-Suppl. 1):133.
• Hussein, M., K. J. Harvatine, Y. R. Boisclair, and D. E. Bauman. 2011. Effect of fish oil supplementation on lipogenic gene expression in mammary gland and liver of mice. FASEB J 25:599.7

Progress 04/15/10 to 04/14/11

Outputs
OUTPUTS: Milk is an important source of nutrients and fat is the most variable milk component. Our current objective is to investigate the mechanism by which bioactive fatty acids reduce milk fat. We have tested the functional role of two candidate genes, Sterol Response Element Binding Protein 1 (SREBP1) and Thyroid Hormone Responsive Spot 14 (S14), in the inhibition of milk fat synthesis using transgenic mouse models. We have determined that SREBP1 and S14 are essential to regulation of lipid synthesis by trans-10,cis-12 conjugated linoleic acid in the mammary gland. In addition, we have designed promoter specific quantitative Real-Time PCR assays for splice variants of acetyl-CoA carboxylase, a key enzyme in fat synthesis. We have also determined that chemical activation of endoplasmic reticulum stress does not reduce fat synthesis in mammary epithelial cells and that inhibition of ER stress does not block trans-10,cis-12 CLA inhibition of lipogenesis. During the first year of the grant we have published two invited review article, two peer reviewed papers, presented four abstracts at national meetings, and spoke at industry conferences. In summary, we have met key objectives of determining the functional role of SREBP1, S14, and ER stress in biohydrogenation intermediate inhibition of milk fat synthesis. Experiments in other objectives have also been initiated. PARTICIPANTS: Nothing significant to report during this reporting period. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Milk is an economically and nutritionally important component of milk, but is very responsive to diet. We have determined the functional role of two candidate regulatory systems, SREBP1 and S14, in diet inhibition of milk fat synthesis and have determined that a third candidate regulatory system (ER stress) appears to not be directly involved. Understanding the functional role of regulatory systems in the dietary regulation of milk fat synthesis is essential to development of targeted selection and management systems to increase milk fat.

Publications

• Bauman, D. E., K. J. Harvatine, and A. L. Lock. 2010. Nutrigenomics, rumen-derived bioactive fatty acids, and the regulation of milk fat synthesis. An. Rev. Nutr. (In Press).
• Foote, M. R., S. L. Giesy, G. Bernal-Santos, D. E. Bauman, and Y. R. Boisclair. 2010. t10,c12-CLA decreases adiposity in peripubertal mice without dose-related detrimental effects on mammary development, inflammation status and metabolism. Am. J. Physiol. Regul. Integr. Comp. Physiol. 299:R1521-28.
• Rudolph, M. C., J. Monks, V. Burns, M. Phistry, R. Marians, M. R. Foote, D. E. Bauman, S. M. Anderson, and M. C. Neville. 2010. Sterol Regulatory Element Binding Protein (Srebf-1) and Dietary Lipid Regulation of Fatty Acid Synthesis in the Mammary Epithelium. Am. J. Physiol. Endocrinol. Metab. 299: E918-27.
• Bauman, D. E., M. A. McGuire, and K. J. Harvatine. 2010. Milk fat Biosynthesis and Secretion: Milk Fat. Encyl Animal Sci. 3:352-8.