The bZIP-like motif of hnRNP C directs the nuclear accumulation of pre-mRNA and lethality in yeast

The hnRNP C protein tetramer cooperatively binds 230 nt increments of pre-m RNA in vitro in a salt-resistant manner and is located along the length of vertebrate transcripts in vivo. Based on these and other findings it has be en suggested that hnRNP C functions as a chaperonin to maintain long length s of RNA topologically single-stranded and accessible to splicing factors. We report here that human C protein is lethal when expressed in the yeast S accharomyces cerevisiae. Through a series of fluorescent immunolocalization studies, lethality was observed to be associated with the rapid nuclear ac cumulation of both C protein and yeast pre-mRNA. Studies using various prot ein constructs and the two hybrid assay reveal that these events are depend ent on the basic 40 residue high-affinity RNA binding domain and its contig uous leucine zipper-like motif (the bZLM, residues 140-214). Additionally, equilibrium binding studies have shown that the bZLM is the determinant of C protein's salt-resistant RNA binding mode. Taken together, these findings further distinguish the bZIP-like domain as the major determinant of C pro tein's high-affinity interaction with RNA, oligomerization, and its highly cooperative RNA binding activity. Finally, these findings indicate that yea st and vertebrates may possess a conserved mechanism for general import of RNP although a true homolog to vertebrate C protein appears not to exist in yeast. Lethality is likely due to the absence in yeast of specific mechani sms for the removal of human C protein from nascent transcripts. (C) 2001 A cademic Press.