|Cross, Brandie -|
|Hack, Anniesha -|
|Rao, Rajini -|
Submitted to: PLoS One
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 20, 2013
Publication Date: June 28, 2013
Repository URL: http://handle.nal.usda.gov/10113/57302
Citation: Cross, B.M., Hack, A., Reinhardt, T.A., Rao, R. 2013. SPCA2 regulates Orai1 trafficking and store independent Ca2+ entry in a model of lactation. PLoS One. 8(6):e67348. Interpretive Summary: A significant and economically important clinical/subclinical metabolic disease in dairy cattle is milk fever. Milk fever and subclinical milk fever increases the incidence of secondary diseases in transition dairy cows that have adverse economic impacts on dairy production profitability. We know that the calcium drain from the body to the mammary gland is a primary key to calcium loss and milk fever development. However we have not known what cellular machinery was controlling this calcium loss. This research paper, through a series of experiments, identifies 2 key proteins that are responsible for baso-lateral calcium transport into the mammary gland and thus loss of calcium from the body stores. The first protein is named Orai1, which is a calcium channel and it combines with the second protein SPCA2 that is a calcium pump. Together they regulate, what we now call, Store Independent Ca2+ Entry (SICE) to support lactation calcium needs. Now that the members of this key lactation calcium pathway have been identified we may be able, at a future date, to regulate their function to prevent milk fever development and thus save dairy farmers profits.
Technical Abstract: An unconventional interaction between SPCA2, an isoform of the Golgi secretory pathway Ca2+-ATPase, and the Ca2+ influx channel Orai1 has previously been shown to contribute to elevated Ca2+ influx in breast cancer derived cells. In order to investigate the physiological role of this interaction, we examined expression and localization of SPCA2 and Orai1 in mouse lactating mammary glands. We observed co-induction and co-immunoprecipitation of both proteins, and isoform-specific differences in the localization of SPCA1 and SPCA2. Three-dimensional cultures of mouse mammary epithelial cells were established using lactogenic hormones and basement membrane. These mammospheres displayed elevated Ca2+ influx by store independent mechanisms, consistent with upregulation of both SPCA2 and Orai1. Knockdown of either SPCA2 or Orai1 severely depleted Ca2+ influx and interfered with mammosphere differentiation. Unexpectedly, knockdown of SPCA2 also inhibited store-dependent mechanism of Ca2+ entry. We show that SPCA2 is required for plasma membrane trafficking of Orai1 in mouse mammary epithelial cells and that this function can be replaced, at least in part, by a membrane anchored C-terminal domain of SPCA2. Our findings reveal a critical role for SPCA2 in eliciting basolateral Ca2+ influx in lactating mammary epithelia by interaction with Orai1.