IDENTIFICATION OF NOVEL ISOFORMS OF THE DELTA-SUBUNIT OF CA2-DEPENDENT PROTEIN KINASE-II - DIFFERENTIAL EXPRESSION IN RAT-BRAIN AND AORTA( CALMODULIN)

Two novel isoforms of the Ca2+/calmodulin-dependent protein kinase II delta subunit were detected in rat aorta. Identification of the subuni ts was based on two independent lines of evidence, i.e. detection by i mmunoblotting of differently sized delta subunits and DNA sequence ana lysis of partial cDNA clones of the kinase. Cytosolic extracts from ra t brain, aorta, and cultured aortic cells were analyzed by Western blo tting using a delta subunit-specific antipeptide antibody. Aortic extr acts demonstrated a single 53-kDa cross-reactive band approximately 7 kDa smaller than the cross-reactive band seen in brain. To ascertain t he structural basis for this difference, reverse-transcribed RNAs from rat aorta and brain were analyzed by polymerase chain reaction (PCR), and the PCR fragments were cloned and sequenced. When aortic cDNA was analyzed with a primer pair that spanned the known variable region of the brain kinase subunit, the amplified PCR products were smaller tha n the major product obtained from brain cDNA. The aortic PCR product w as cloned and sequenced and found to represent two novel subunit seque nces, designated delta2 and delta3 to distinguish them from the previo usly described delta sequence (now called delta1) from brain. Delta2 w as identical to the predicted delta1 sequence except for a deletion of 102 base pairs (bp). This deletion corresponded to nearly the entire variable domain. In the sequence of delta3, this 102-bp region was rep laced by a sequence of 33 bp that had 79% nucleotide sequence identity to a portion of the gamma subunit variable domain. A fourth form of t he delta subunit (delta4) was identified in rat skeletal muscle. The d elta4 isoform was characterized by the deletion of a 42-bp sequence id entical to the 42 bp at the 3' end of the 102-bp deletion of delta2. R everse-transcription PCR analysis of additional rat tissues indicated that alternatively spliced variants of the delta subunit of Ca2+/calmo dulin-dependent protein kinase II are expressed in a tissue-specific p attern.