Cloning a novel MAL protein from pancreatic Î²-cells
A small gene family may be involved in sorting proteins to rafts and hence transport to intracellular organelles and the plasma membrane. MAL is found to localise to lipid rafts and is involved in the sorting of apical proteins in polarised epithelial cell types. BENE and Plasmolipin have also been associated with lipid rafts in either endothelial or glial cells. However, MAL, BENE and Plasmolipin are not expressed in endocrine tissues. Recently, a human EST of partial sequence was proposed to be the fourth member of this family. During the current study, the complete coding sequence of this member, termed MAL2, was identified in rat b-cells of the pancreas. Furthermore, through library screening and 5’ RACE, two isoforms of MAL2 were identified - rMAL2-A and rMAL2-B. The two isoforms differ at their amino termini and rMAL2-B contains a long 5’ UTR with upstream AUG (uAUG) codons and two open reading frames. Analysis of these uAUG codons found they were capable of decreasing expression of a reporter gene construct and likely to contribute to the observed low expression of rMAL2-B. Both isoforms were found to be glycosylated on the second extracellular/intralumenal loop. rMAL2-A and rMAL2-B appeared not to display raft association within b-cells, although further work is required to refine the extraction procedure. Subcellular analysis of the two isoforms found that they mainly localised to the Golgi, overlapping in distribution with TGN38, an endogenous TGN marker.