Enzymatic activity has been identified in cultured cells which catalyzes the removal of intact mono(ADP-ribose) residues which are bound to protein at arginine. Other activities have been detected which catalyze the removal of ADP-ribose via the sequential removal of AMP and ribose-5-phosphate.

The objectives of these studies were to assess U.S. dairy nutritionists' approaches toward balancing CP in lactating cow diets, and to leverage existing knowledge of postabsorptive AA metabolism through the application of a mathematical ration-balancing model to predict N efficiency through a more accurate postabsorptive amino acid (AA) delivery. In experiment 1, dairy nutritionists (n = 77) that fed a total of 521,000 lactating dairy cows responded to a questionnaire related to demographic information, feelings toward environmental nitrogen (N) excretion, and dietary CP balancing practices. Eighty-nine percent of nutritionists balanced diets based on one or more individual AA requirements of dairy cows. The primary concern with formulation of lower CP diets was the cost per unit of metabolizable protein (MP). In the second study, three treatments were fed to lactating Holstein cows (n = 48) to test proof of concept of NASEM 2021 and a nonlinear optimizer: a control balanced to fulfill all nutrient needs of lactating dairy cows producing 45 kg milk/d using the NRC (2001) dairy model (NRC01), and two diets balanced with a nonlinear optimizer that fulfilled requirements according to the updated NASEM (2021) dairy model and attempted to either: 1) maximize N efficiency through tailored post-ruminal AA supply (NEFF), or 2) maximize income over feed cost (IOFC). A simulation function was written in RStudio (version 2022.02.3) to predict daily animal performance with NASEM 2021. Dry matter intake, milk, and milk components from both the observed data and the simulation data matrices were analyzed as repeated measures (days) in a mixed model to test for both observed and predicted (simulated) differences in treatment means. Income over feed cost was $4.83, $4.77, and $5.12/cow/d for NRC01, IOFC, and NEFF, respectively (P = 0.96). Nitrogen efficiency (%) was greatest for NEFF (33.7), followed by NRC01 (28.9) and IOFC (23.4; P < 0.05 between all treatments). Based upon simulation data, NASEM 2021 predicted relative performance differences between animals that consumed treatments with differing absorbed EAA supplies, although residual analyses revealed that further progress could be made in milk protein (g/d), milk fat (g/d), milk yield (kg/d), and DMI (kg/d) predictions. / Doctor of Philosophy / Nitrogen (N) is considered one of the major environmental pollutants of the dairy industry. A concerted focus on reducing dietary crude protein (CP) loss by targeting the specific amino acid (AA) needs of cattle will also reduce the amount of N they excrete. Because most dairy farmers hire nutritionists to formulate rations for their cows, the objective of our first study, performed in 2019, was to assess U.S. dairy nutritionists' approaches toward balancing AA in lactating cow diets. The objective of our second study was to apply a mathematical ration-balancing model that specifically optimized AA efficiency for lactating dairy cows by leveraging existing knowledge of dairy cow AA metabolism. In study 1, 89% of nutritionists balanced diets based on one or more individual AA requirements of dairy cows. Most respondents (72%) reported that they are currently formulating diets with lower dietary protein than they were 3 to 5 yr ago. In the first study, the primary concern with formulating lower protein diets was the high cost of protein sources that are highly metabolizable for cows. In study 2, income over feed cost (income made from estimated milk sales minus cost of feed) was similar for cows fed a diet with the lowest protein (relative to two other diets), but cows on the low protein diet also ate the least (kg of dry matter/d) and produced the least amount of milk protein (kg/d) and milk fat (kg/d). Still, relative to the other two diets in the study, N consumed by cows on the lowest CP diet was more efficiently converted to milk protein, which resulted in lesser N excretion in milk. Harnessing the knowledge gained from these studies could allow significant reduction of environmental N excretion at little to no cost to consumers or farmers.

Chronic subclinical levels of disease occur frequently in intensive swine production and compromise nutrient utilization efficiency. Sulfur amino acids (methionine plus cysteine; M+C) have been implicated in improving the animal’s response to immune system stimulation (ISS). Research objectives were to determine the effects of ISS on the optimal dietary methionine to methionine plus cysteine ratio (M:M+C) and on the fractional synthesis rate (FSR) of albumin, fibrinogen and total protein in plasma, liver, and small intestine (SI) of growing pigs. A nitrogen balance study showed that the optimal M:M+C was increased during ISS and greater than 0.62. In a flooding dose infusion study it was determined that total plasma protein FSR was increased during ISS and tended to decrease with reduced M+C intake. Plasma albumin FSR decreased with reduced M+C intake. The data implicates M+C as important nutrients involved in the immune response and careful dietary supplementation during ISS is necessary. / Funding sponsored by Evonik Degussa, Ontario Pork, the Ontario Ministry of Agriculture, Food and Rural Affairs and the Natural Sciences and Engineering Research Council of Canada.

This experiment was conducted to determine if changing the amount of ideal aminoacids (meaning the amount of amino acids necessary to supply all the animal's needswithout excesses or deficiencies of any single amino acid), in relation to caloric intakewill change nitrogen metabolism and weight loss in obese mature dogs. Informationprovided by this experiment can be used to formulate canine diets emphasizing weightloss in older animals.Six moderately obese mature female crossbred hounds were fed diets varying in theirratio of lysine:calories (Lysine % : Mcal ME/g) (2.2, 3.0, and 3.8) in a 3 x 3 replicatedLatin square design. Increasing the lysine:calorie of the diets linearly increased theamount of nitrogen absorbed. It did not, however, significantly affect blood chemistryvalues. Protein turnover exhibited a positive linear trend with increasing ratio and proteindegradation showed a strong quadratic change with the lowest point of degradationoccurring with the diet containing a 3.0 lysine:calorie ratio. Plasma urea and creatinineexcretion demonstrated quadratic tendencies with the two highest values occurring withthe diets containing lysine:calorie of 2.2 and 3.8, reflecting changes in muscle proteinbreakdown while nitrogen was retained in the body. Caloric restriction did not result inloss of lean mass as much as a loss of fat mass. By increasing the quality of protein fedas a percentage of caloric intake, lean muscle mass was conserved during periods ofcaloric restriction.

Equine pituitary pars intermedia dysfunction (PPID) typically occurs in horses older than 15 years of age and is characterized by hair coat abnormalities, muscle atrophy and decreased insulin sensitivity. The first objective of this research was to compare the rate of whole body protein metabolism and relative abundance of key factors in the signaling pathways associated with muscle protein synthesis and protein breakdown in response to feeding in Control and PPID horses. No differences (P > 0.05) were seen between the PPID and Control groups in whole-body protein metabolism or post-prandial activation of the muscle signaling pathways regulating skeletal muscle protein synthesis and breakdown. The second objective of this research was to determine if aged horses with PPID had reduced insulin sensitivity and alterations in the insulin-mediated signaling pathways in the skeletal muscle when compared to non-PPID, aged Control horses. Measures of insulin sensitivity and the activation of factors associated with protein synthesis and breakdown were similar between the PPID and Control groups (P > 0.05). Overall, insulin sensitivity and protein metabolism are similar between the PPID and Control groups. The studies suggest that abnormalities may exist as a function of advanced age rather than PPID status directly.

xvi, 227 leaves : ill. ; 26 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Physiology, 1971

Typescript (photocopy) / xi, 217 leaves ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Agronomy, 1980

Les mécanismes d’adaptation environnementale du métabolisme protéique à court et long-terme sont mal connus. L’exposition prolongée à la chaleur (32°C vs. 22°C) modifie l'expression de quelques gènes impliqués dans le métabolisme protéino-énergétique du muscle Pectoralis major de poulet. La moindre activation de la protéine ribosomale S6 par des facteurs anaboliques à 32°C pourrait indiquer une baisse de l’efficacité de la traduction des ARNm en protéines au chaud. Ceci, associé à une baisse du potentiel de synthèse, traduit une altération à long-terme de la protéosynthèse musculaire au chaud. La distribution séquentielle de régimes alimentaires variant par leur teneur en protéines et/ou en énergie induit au niveau du muscle Pectoralis major de poulet des régulations considérables d’acteurs impliqués dans le contrôle de la protéolyse (e.g. atrogin-1) et de la protéosynthèse (e.g. mTOR, S6K1, S6). Ceci indique une régulation à court-terme de l’équilibre protéosynthèse/protéolyse, dont les mécanismes et les limites restent à caractériser. / Molecular mechanisms underlying the short and long-term environmental adaptation of protein metabolism are not well understood. Prolonged heat exposure (32°C vs. 22°C) modified the expression of some genes related to protein and energy metabolism in the Pectoralis major muscle of chickens. The lower activation of the ribosomal protein S6 by anabolic factors at 32°C could indicate a decrease in the efficiency of mRNA translation into proteins in hot environment. These findings, associated with an impaired potential of protein synthesis, suggest a long-term alteration of muscle protein synthesis under heat conditions. Sequential distribution of diets varying in protein and/or energy contents induced drastic regulations of genes and proteins involved in the control of proteolysis (e.g. atrogin-1) and protein synthesis (e.g. mTOR, S6K1, S6) in the Pectoralis major muscle of chickens. This may indicate a short-term regulation of protein synthesis/proteolysis balance, whose mechanisms and limits remain to be characterized.

Orientador: Maria Cristina Cintra Gomes Marcondes / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-26T13:25:51Z (GMT). No. of bitstreams: 1 Cruz_BreadLeandroGomesda_D.pdf: 1846989 bytes, checksum: 30a328ba2d64a0e66c6bb74d4c81d492 (MD5) Previous issue date: 2014 / Resumo: O câncer é uma das principais causas de morte no mundo e o quadro de caquexia provocado por alguns tipos de tumor é um dos grandes responsáveis por isso. A caquexia instaurada em pacientes com câncer, sendo mais prevalente nos tumores gastrointestinal, pulmonar, pancreático e mamário, é caracterizada, dentre outros processos, pela perda involuntária de peso, devido a constante espoliação sobre a massa magra corporal. Estudos que tenham como objetivo a manutenção da massa magra em organismos portadores de tumor caquético são importantes para contribuir com a redução de óbitos e preservar a qualidade de vida das pessoas com câncer. Nos últimos anos, a leucina tem mostrado ser eficaz na manutenção da massa magra corpórea através do estímulo de síntese protéica muscular e inibição da degradação de proteína, principalmente da massa magra corpórea. Logo, entender como a presença da leucina estimula a síntese protéica e atua de forma protetora em organismo no estado caquético tem se mostrado uma necessidade crescente. Desse modo, a proposta deste trabalho foi avaliar, ao longo do tempo, os efeitos de dieta rica em leucina sobre a sinalização de síntese e degradação protéica, envolvendo o complexo mTOR em músculos de ratos portadores do carcinossarcoma de Walker 256. Os animais foram distribuídos em grupos de acordo com a inoculação tumoral e/ou esquema nutricional com dieta rica em leucina, sendo sacrificados em três momentos diferentes do desenvolvimento tumoral (7º, 14º e 21º dias após a implantação do tumor). No músculo gastrocnêmio foram analisadas as proteínas-chave da via mTOR, como RAG A GTPase, ERK/MAP4K3, PKB/Akt, mTOR, p70S6K1, Jnk, IRS-1, STAT3, e STAT6 e, tambem, foram avaliadas as proteínas de degradação protéica pertencentes ao sistema ubiquitina-proteossomo (11S, 19S e 20S) e citocinas IL-4, IL-6, IL-10, TNF? e INF?. Os resultados mostram que o desenvolvimento tumoral reduziu a ativação de RAG-A, associada com queda de IRS-1, aumento da PKB/Akt e Erk/MAP4K3 no 21º dia e manutenção de p70S6K1; também houve aumento dos níveis de STAT-3 e STAT-6 em ratos portadores de tumor. Entretanto, a presença de leucina na dieta modulou etapas chave da via mTOR pelo desencadeamento da ativação aumentada de RAG-A e mTOR junto com a manutenção dos níveis de JNK, STAT-3 e STAT-6 no músculo durante o desenvolvimento do tumor de Walker no organismo caquético. A análise da sinalização para degradação protéica mostrou que o crescimento tumoral promoveu, simultaneamente, diminuição de proteína muscular, acentuado aumento de citocinas pró-inflamatórias (TNF?, IL-6 e IFN?) e aumento progressivo das subunidades proteossômicas (19S e 20S), sendo que a suplementação com leucina atenuou essa ativação. Os resultados obtidos apoiam o efeito benéfico do uso de leucina e esclarece as vias metabólicas utilizadas por este aminoácido, contribuindo para melhor compreensão da ação in vivo desse aminoácido sobre a caquexia / Abstract: Cancer is one of the most important causes of death worldwide and the process of cachexia caused by some types of tumour is largely responsible for this. Cancer-cachexia state established in these patients is characterized, among other processes, by involuntary loss of weight due to constant spoliation of lean body mass. Many studies aim to focus in maintenance of lean body mass in cachectic tumour-bearing host, contributing to the reduction of deaths and improving the quality of life of cancer patients. In recent years, leucine has been shown to be effective in maintaining lean body mass by stimulating muscle protein synthesis and inhibiting the proteolysis. Therefore, there are increased needs in understanding how the presence of leucine stimulates protein synthesis and acts protectively in the cachectic state. The aim of this work is to evaluate the effects of leucine-rich diet in a time-course model on signalling protein synthesis and degradation involving mTOR complex in muscles of Walker 256 carcinoma-bearing rats. Animals were divided into experimental groups based on the tumour inoculation and/or fed a nutritional supplementation diet rich in leucine. Animals were sacrificed at three different times depending on the tumour development (7, 14 and 21 days after tumour implantation), and the gastrocnemius muscles were analysed as mTOR pathway and ubiquitin-proteasome via. The results showed that the tumour development has reduced the activation of RAG-A, associated with a decrease of IRS-1, increased PKB / Akt and Erk / MAP4K3 at day 21 and maintaining p70S6K1, there has also been increasing levels of STAT-3 and STAT-6 in tumour-bearing rats. Meanwhile, the presence of leucine in the diet modulated the key steps of the mTOR pathway for triggering the increased RAG-A and mTOR activation along with the maintenance of levels of JNK, STAT-3 and STAT-6 in the muscle during tumour development in cachectic host. The gastrocnemius muscle signalling of protein degradation indicated by the ubiquitin-proteasome subunits (11S, 19S and 20S) and pro- and anti-inflammatory cytokines were marked increase of pro-inflammatory cytokines (TNF?, IL-6 and IFN?) and a progressive increase in the proteasome subunits (19S and 20S) associated with simultaneously decreased muscle protein. The supplementation with leucine attenuated these parameters, suggesting the beneficial effect of the use of leucine and clarifies the metabolic pathways used by this amino acid, contributing to better understanding of the in vivo action of this amino acid on cachexia / Doutorado / Fisiologia / Doutor em Biologia Funcional e Molecular

The cytoskeleton is an intrinsic aspect of all cells, and is essential for many cellular events including cell motility, endocytosis, cell division and wound healing. Remodeling of the cytoskeleton in response to these cellular activities leads to significant alterations in the morphology of the cell. One such alteration is the formation of an actomyosin contractile array required for cytokinesis, wound healing and embryonic development. / Cellular structure and shape depends upon tensional prestress brought about by the organization of cytoskeletal components. Using the Xenopus laevis oocyte wound healing model, it is first described how diminished cellular tension affects the balance of the Rho family of GTPases, and subsequently prevents the formation of actomyosin contractile arrays. This suggests that cellular tension in the cell is not created at the level of the cytoskeletal elements but rather via the upstream signaling molecules: RhoA and Cdc42. / The role of N-WASP (Neural-Wiscott Aldrich Syndrome Protein), a mediator of Arp2/3 based actin polymerization, is next examined for its putative role in cellular wound healing. Xenopus laevis oocytes injected with mutant N-WASP constructs reveals in vivo evidence that functional N-WASP is required for appropriate contractile array formation and wound closure. / Lastly, it is revealed that the cellular structures involved with single cell wound healing in other model systems are also important for the initial repair of severed muscle cells. Actin, non-muscle myosin-II, microtubules, sarcomeric myosin and Cdc42 are all recruited and reorganized at the edge of damaged C2C12 myotubes. This data promotes the possibility that an actomyosin array may be established in injured muscle cells as well.