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Casein kinase activity in rat mammary gland Golgi vesicles. Demonstration of latency and requirement for a transmembrane ATP carrier.

ABSTRACT: A Golgi vesicle-enriched preparation from mammary tissue of lactating rats has been used to investigate the phosphorylation of caseins in vitro. Casein kinase, together with its casein substrates, is enclosed within the lumen of Golgi membrane vesicles and has a requirement for Ca2+ and ATP. The permeability characteristics of the Golgi membrane to ATP and Ca2+ therefore have a possible regulatory influence on casein kinase activity. This influence has been investigated by alteration of the permeability characteristics by using several agents having differing degrees of selectivity. The ionophore A23187, which permits loss of Ca2+ from the vesicles, caused a decrease in casein phosphorylation which could be reversed by externally supplied Ca2+. Alamethicin, an ionophore that creates larger transmembrane channels, caused an increase in casein phosphorylation. This increase showed a requirement for divalent metal ions which could be satisfied by either Ca2+ or Mn2+. Under the same conditions, La3+ was inhibitory. Triton X-100 caused loss of intravesicular Ca2+, yet this was accompanied by an increase in phosphate incorporation into the caseins. We conclude from these results that the binding site on casein kinase for ATP is within the Golgi membrane barrier and that they imply the presence of a transmembrane ATP-transport mechanism. Inhibition of casein phosphorylation by atractyloside and carboxyatractyloside lends support to this concept.

SUBMITTER: West DW

PROVIDER: S-EPMC1153463 | biostudies-other | 1984 Apr

REPOSITORIES: biostudies-other

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Project description:BACKGROUND:Mounting evidences observed that subacute ruminal acidosis (SARA) induced by high concentration (HC) diet increases the translocation of histamine from digestive tract into circulation causing a diverse of diseases in dairy cows. However, it is largely unknown how it does affect the function of mammary gland and milk quality. Hence, this study aims to observe the effects of histamine derived from the digestive tract on the inflammatory response and casein synthesis in the mammary glands during SARA. Twelve cows fitted rumen fistula were randomly divided into either control group administrated low concentration (LC) diet (60% forage, n?=?6) or treatment group administrated HC diet (40% forage, n?=?6) for 18 weeks. RESULTS:Our data showed that HC diet resulted in significant declines in rumen pH value, milk yield and milk quality, as well as longer duration of averaged pH value below 5.6 per day (more than 180 min) compared to LC diet, these findings confirmed SARA occurence. Our study also observed that SARA increased the content of histamine in rumen fluid, plasma, liver and mammary gland, and enhanced the mRNA expression of histamine specific receptor in the mammary gland. Additionally, we found that the mRNA expression of inflammatory response genes in mammary glands was increased, which was consistent with the protein expression results, showing that the protein kinase C(PKC) / nuclear factor kappa B (NF-?B) or protein kinase A (PKA) / NF-?B signalling pathways of the inflammatory response were activated. The mRNA expression of mTOR, P70S6K and ?S1 in mammary glands were significantly decreased with the protein expression of mTOR, P70S6K and ?S1-casein, and the phosphorylation levels of the mTOR and P70S6K proteins were also decreased. CONCLUSIONS:Our study showed that the milk protein of lactating cows is depressed after long-term feeding of HC at the individual level, which was paralleled at the gene and protein levels. The inflammatory response in mammary gland caused by histamine derived from the digestive tract is related to the decline of casein synthesis. Our findings point to a new link between the inflammatory response and casein synthesis, but the understanding of the molecular mechanisms involved in this process will require further research.

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Casein kinase activity in rat mammary gland Golgi vesicles. Demonstration of latency and requirement for a transmembrane ATP carrier.

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