Cloning and sequencing of a human cDNA from cytolytic NK/T cells with homology to yeast SEC7

Abstract

Using the technique of subtractive hybridization (natural killer (NK) minus T helper cell line Jurkat), a human cDNA clone named B2-1 was isolated and sequenced. B2-1 transcripts are highly expressed in NK and peripheral T cells, but not in the T helper cell line used in the subtraction. Two overlapping cDNA clones contained 3309 bp, in agreement with a predicted size of 3.2 kb obtained by Northern blot analysis. The cDNA contained a 5' open reading frame encoding a 398 amino acid (aa) protein with a calculated M(r) of 46,383. The protein contains N-linked glycosylation sites and a site for phosphorylation by protein kinase C. Database homology searches show that the deduced protein is novel and has three discrete domains with significant homology to several structural proteins. The most striking homology was to yeast SEC7 in the central domain of the gene (57% identical over 466 bp) and also the protein level (42% identical amino acids; 39% conserved amino acids). SEC7 is a large protein (2008 amino acids) found on Golgi vesicles that plays a role in protein transport.