Alternative splicing generates a smaller assortment of Ca_V2.1 transcripts in cerebellar Purkinje cells than in the cerebellum

Abstract

Alternative splicing generates a smaller assortment of CaV2.1 transcripts in cerebellar Purkinje cells than in the cerebellum. Physiol Genomics 24: 86 -96, 2006. First published November 8, 2005 doi:10.1152/physiolgenomics.00149.2005.-P/Qtype calcium channels control many calcium-driven functions in the brain. The CACNA1A gene encoding the pore-forming CaV2.1 (␣1A) subunit of P/Q-type channels undergoes alternative splicing at multiple loci. This results in channel variants with different phenotypes. However, the combinatorial patterns of alternative splice events at two or more loci, and hence the diversity of CaV2.1 transcripts, are incompletely defined for specific brain regions and types of brain neurons. Using RT-PCR and splice variant-specific primers, we have identified multiple CaV2.1 transcript variants defined by different pairs of splice events in the cerebellum of adult rat. We have uncovered new splice variations between exons 28 and 34 (some of which predict a premature stop codon) and a new variation in exon 47 (which predicts a novel extended COOH-terminus). Single cell RT-PCR reveals that each individual cerebellar Purkinje neuron also expresses multiple alternative CaV2.1 transcripts, but the assortment is smaller than in the cerebellum. Two of these variants encode different extended COOH-termini which are not the same as those previously reported in Purkinje cells of the mouse. Our patch-clamp recordings show that calcium channel currents in the soma and dendrites of Purkinje cells are largely inhibited by a concentration of -agatoxin IVA selective for P-type over Q-type channels, suggesting that the different transcripts may form phenotypic variants of P-type calcium channels in Purkinje cells. These results expand the known diversity of Ca V2.1 transcripts in cerebellar Purkinje cells, and propose the selective expression of distinct assortments of Ca V2.1 transcripts in different brain neurons and species. splice variants; P type; calcium channels; Purkinje neurons 0 VOLTAGE-GATED calcium (Ca 2ϩ ) channels regulate numerous cellular functions, from neuronal electrical activity to intracellular signaling pathways. Diversity of Ca 2ϩ channels in the brain is apparent from the many roles that Ca 2ϩ channels play in different cell types and subcellular compartments (3). The diversity reflects, in part, the existence of nine classes of the pore-forming Ca V (␣1) subunit in the brain and alternative pre-mRNA splicing of the gene encoding each class of Ca V subunit (12, 17). P/Q-type Ca 2ϩ channels control many functions throughout the brain and contain a Ca V 2.1 subunit (also known as ␣ 1A ). However, information about splicing of the CACNA1A gene encoding Ca V 2.1 in different brain regions and neurons is incomplete (2,5,16,30,33,35,37). Augmentation of what is currently known about Ca V 2.1 splicing in the cerebellum and cerebellar Purkinje cells is of particular interest, because P-type currents in Purkinje cells are the prototypical P-type currents against which putative P-type and Q-...

Alternative splicing regulates activation-induced cytidine deaminase (AID): implications for suppression of AID mutagenic activity in normal and malignant B cells

“…Finally, our data demonstrating that AID is expressed in a "one variant per cell" manner raises the question of whether this occurs more frequently in nature, and perhaps has been largely missed because traditional analysis of alternative splicing is performed using total RNA from bulky cell populations or tissue. To our knowledge, the only other reports of alternatively spliced genes expressed in a "one variant per cell" manner are prepro-tachykinin A 51 , the calcium channel protein CaV2.1, 52 and SCN5A. 53 These 3 reports, however, did not discuss the implications of their results from the standpoint of regulation of alternatively spliced genes.…”

CaV2.1 voltage activated calcium channels and synaptic transmission in familial hemiplegic migraine pathogenesis

“…Cav2.1 channels are located throughout the mammalian brain and spinal cord (Westenbroek et al 1995) at presynaptic terminals (Catterall 1998, Wu et al 1999, Mintz et al 1995 and at somatodendritic membranes. The CACNA1A gene encoding Cav2.1 channels undergoes alternative splicing at multiple loci in an age-, gender-, and species-dependent manner (Bourinet et al 1999, Chang et al 2007, Chaudhuri et al 2005, Kanumilli et al 2006, Soong et al 2002. This mechanism resultes in multiple Cav2.1 splice variants with different outcomes in neuronal distribution and subcellular localization, biophysical properties, and sensitivity to ω-AgaIVA.…”

Old proteins, developing roles: The regulation of calcium channels by synaptic proteins

“…In addition to a number of Ca V 2.1 and Ca V 2.2 splice variants that encode variations outside of the major synaptic protein interaction regions that show altered biophysical properties, [134][135][136] alternate splicing of the II-III linkers and C-termini, notably by exons e18a or e37a and b respectively can add further functional diversity (Table 2). 137 P/Q-type splice variants lacking parts of the synprint region showed reduced current and a large (40 mV) rightward shift in inactivation, 138 while N-type variants with substantial deletions of the synprint motif show decreased ω-conotoxin MVIIA and GVIA sensitivity, depolarized voltage dependence of steady-state inactivation, and enhanced recovery from inactivation.…”