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Characterization of the Skeletal Muscle Calpain/Calpastatin System in Growth Models in Swine and Cattle" (2013)

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Characterization of the Skeletal Muscle Calpain/Calpastatin System in Growth Models in Swine and Cattle Iowa State University Capstones, Theses and Graduate Theses and Dissertations Dissertations 2013 Characterization of the skeletal muscle calpain/ calpastatin system in growth models in swine and cattle Shannon Michelle Cruzen Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/etd Part of the Agriculture Commons, Animal Sciences Commons, and the Food Science Commons Recommended Citation Cruzen, Shannon Michelle, "Characterization of the skeletal muscle calpain/calpastatin system in growth models in swine and cattle" (2013). Graduate Theses and Dissertations. 13305. https://lib.dr.iastate.edu/etd/13305 This Dissertation is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Characterization of the skeletal muscle calpain/calpastatin system in growth models in swine and cattle by Shannon Michelle Cruzen A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Meat Science Program of Study Committee: Elisabeth Huff-Lonergan, Major Professor Steven Lonergan Philip Dixon Ted Huiatt Joseph Sebranek Iowa State University Ames, Iowa 2013 Copyright © Shannon Michelle Cruzen, 2013. All rights reserved. ii TABLE OF CONTENTS Page ACKNOWLEDGEMENTS ....................................................................................... iv ABSTRACT ............................................................................................................ vi CHAPTER 1 GENERAL INTRODUCTION .......................................................... 1 Efficiency ...................................................................................................... 2 Growth .......................................................................................................... 2 Further Characterization ............................................................................... 3 Dissertation Organization .............................................................................. 4 Literature Cited .............................................................................................. 5 CHAPTER 2 REVIEW OF LITERATURE ............................................................ 7 Introduction .................................................................................................... 7 Muscle Protein Degradation and Synthesis .................................................. 10 Efficiency and Residual Feed Intake ............................................................ 41 Animal Age and Meat Quality ....................................................................... 47 Conclusion .................................................................................................... 51 Literature Cited .............................................................................................. 52 CHAPTER 3 EVIDENCE OF DECREASED MUSCLE PROTEIN TURNOVER IN GILTS SELECTED FOR LOW RESIDUAL FEED INTAKE .......................... 73 Abstract .......................................................................................................... 73 Introduction .................................................................................................... 74 Materials and Methods ................................................................................... 75 Results and Discussion ................................................................................. 81 Acknowledgements ....................................................................................... 90 Literature Cited .............................................................................................. 90 Tables and Figures ........................................................................................ 96 CHAPTER 4 POSTMORTEM PROTEOLYSIS IN THREE MUSCLES FROM GROWING AND MATURE BEEF CATTLE.......................................................... 106 Abstract .......................................................................................................... 106 Introduction .................................................................................................... 107 Materials and Methods .................................................................................. 110 Results and Discussion ................................................................................. 115 Conclusions ................................................................................................... 122 iii Page Acknowledgements ....................................................................................... 123 Literature Cited ............................................................................................. 123 Tables and Figures ........................................................................................ 130 CHAPTER 5 SKELETAL MUSCLE CALPASTATINS SEPARATED USING ANION EXCHANGE CHROMATOGRAPHY DIFFER IN PHOSPHORYLATION ............................................................................................. 139 Graphical Abstract ....................................................................................... 139 Abstract ......................................................................................................... 139 Introduction ................................................................................................... 141 Materials and Methods .................................................................................. 146 Results and Discussion ................................................................................. 153 Abbreviations Used ....................................................................................... 160 Acknowledgements ....................................................................................... 161 Literature Cited ............................................................................................. 161 Tables and Figures ........................................................................................ 167 CHAPTER 6 GENERAL DISCUSSION AND CONCLUSIONS .......................... 183 Literature Cited ............................................................................................. 187 iv ACKNOWLEDGEMENTS Completing my PhD would not have been possible without the help and support of everyone around me. I would first like to thank my husband and best friend, Tyler Cruzen, who has been with me every step of the way and joyfully become part of the meat science family, even though all he understands sometimes are the “key words.” Thank you also to my mother, Lori Kiphen, who nurtured my love of learning from as early as I can remember and has always supported me, no matter what I decided to do. I would like to thank my committee chair, Elisabeth Lonergan, for allowing me to work with her. Working with Elisabeth at Iowa State was what I wanted so much that I didn’t have a backup plan, and when I was informed I got the position, my excitement was a sight to behold. I am so grateful for Elisabeth’s respect, guidance, and support throughout these three years. I will always be proud to say I came from the Lonergan Lab, and I hope to work with you again. To Steven Lonergan, thank you for being a mentor and friend, and for working alongside me for hours assaying calpain and calpastatin. Thank you to my other committee members, Joe Sebranek, Philip Dixon, and Ted Huiatt, for their guidance and support throughout the course of this research. In addition, I would also like to thank my friends, colleagues, the department faculty, and staff for making my time at Iowa State University a wonderful experience. Thank you to the graduate and undergraduate students who helped me in the lab, including Emily Arkfeld, Roslyn Punt, and Kristin Adams. Pedro Paulino, thank you for your ideas and support in the lab. I also owe a large debt to Ed Steadham, who kept our lab and my projects running v smoothly and provided endless guidance and wisdom. A huge thanks goes to Kyle Grubbs, who has been my colleague, co-coach, and most importantly, one of my best friends through this process. I think we will be shooting ideas off each other for the rest of our lives. To everyone who has helped me in completing my PhD, this has been an amazing experience, and I am ready to go out and show everyone how great Iowa State University has been. I can only hope to emulate the amazing examples, support systems, and friends that I have had the privilege of interacting with in my time here. vi ABSTRACT The overall objective of the work in this dissertation was to characterize µ-calpain, m-calpain, and calpastatin activities in order to learn more about the system in the live animal as well as its potential effects on postmortem proteolysis. Animal models of efficiency and growth were used to achieve this objective. The first specific objective was to evaluate muscle protein turnover differences in pigs selected for residual feed intake (RFI). Muscle samples were collected, and insulin signaling, protein synthesis, and specific protein degradation proteins were analyzed for expression and activities in twelve gilts each from low (LRFI) and high (HRFI) RFI lines. LRFI (more efficient) pig muscle had greater
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