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-mRNA 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 been suggested that hnRNP C functions as a chaperonin to maintain long lengths of RNA topologically single-stranded and accessible to splicing factors. We report here that human C protein is lethal when expressed in the yeast Saccharomyces cerevisiae. Through a series of fluorescent immunolocalization studies, lethality was observed to be associated with the rapid nuclear accumulation of both C protein and yeast pre-mRNA. Studies using various protein constructs and the two hybrid assay reveal that these events are dependent on the basic 40 residue high-affinity RNA binding domain and its contiguous 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 protein's high-affinity interaction with RNA, oligomerization, and its highly cooperative RNA binding activity. Finally, these findings indicate that yeast 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 mechanisms for the removal of human C protein from nascent transcripts.
Tan
J H
JH
Department of Biological Sciences, Vanderbilt University, Nashville, TN 37235-1634, USA.
Kajiwara
Y
Y
Shahied
L
L
Li
J
J
McAfee
J G
JG
LeStourgeon
W M
WM
D021581:Active Transport, Cell Nucleus; D020816:Amino Acid Motifs; D002467:Cell Nucleus / Q000378:metabolism*; D002942:Circular Dichroism; D005453:Fluorescence; D005804:Genes, Lethal / Q000235:genetics*; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D017403:In Situ Hybridization; D016350:Leucine Zippers; D011485:Protein Binding / Q000187:drug effects; D020836:Protein Structure, Quaternary; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism*; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics* / Q000254:growth & development; D012492:Salts / Q000494:pharmacology; D017384:Sequence Deletion / Q000235:genetics; D013816:Thermodynamics
D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D012322:RNA Precursors; D012331:RNA, Fungal; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012261:Ribonucleoproteins; D012492:Salts