Details

Autor(en) / Beteiligte

• Nourian, Zahra
• Li, Min
• Leo, M. Dennis
• Jaggar, Jonathan H
• Braun, Andrew P
• Hill, Michael A

Titel

Large Conductance Ca.sup.2+-Activated K.sup.+ Channel

Ist Teil von

• PloS one, 2014-06, Vol.9 (6)

Ort / Verlag

Public Library of Science

Erscheinungsjahr

2014

Quelle

EZB Electronic Journals Library

Beschreibungen/Notizen

• Previous studies report functional differences in large conductance Ca.sup.2+ activated-K.sup.+ channels (BK.sub.Ca) of smooth muscle cells (VSMC) from rat cerebral and cremaster muscle resistance arteries. The present studies aimed to determine if this complexity in BK.sub.Ca activity may, in part, be due to splice variants in the pore-forming [alpha]-subunit. BK.sub.Ca variants in the intracellular C terminus of the [alpha]-subunit, and their relative expression to total [alpha]-subunit, were examined by qPCR. Sequencing of RT-PCR products showed two [alpha]-subunit variants, ZERO and STREX, to be identical in cremaster and cerebral arteries. Levels of STREX mRNA expression were, however, significantly higher in cremaster VSMCs (28.9±4.2% of total [alpha]-BK.sub.Ca) compared with cerebral vessels (16.5±0.9%). Further, a low level of BK.sub.Ca SS4 [alpha]-subunit variant was seen in cerebral arteries, while undetectable in cremaster arteries. Protein biotinylation assays, in expression systems and arterial preparations, were used to determine whether differences in splice variant mRNA expression affect surface membrane/cytosolic location of the channel. In AD-293 and CHO-K1 cells, rat STREX was more likely to be located at the plasma membrane compared to ZERO, although the great majority of channel protein was in the membrane in both cases. Co-expression of [beta]1-BK.sub.Ca subunit with STREX or ZERO did not influence the dominant membrane expression of [alpha]-BK.sub.Ca subunits, whereas in the absence of [alpha]-BK.sub.Ca, a significant proportion of [beta]1-subunit remained cytosolic. Biotinylation assays of cremaster and cerebral arteries showed that differences in STREX/ZERO expression do not alter membrane/cytosolic distribution of the channel under basal conditions. These data, however, revealed that the amount of [alpha]-BK.sub.Ca in cerebral arteries is approximately 20X higher than in cremaster vessels. Thus, the data support the major functional differences in BK.sub.Ca activity in cremaster, as compared to cerebral VSMCs, being related to total [alpha]-BK.sub.Ca expression, regardless of differences in splice variant expression.