Binding of LARP6 to the conserved 5' stem-loop regulates translation of mRNAs encoding type I collagen.
Type I collagen is the most abundant protein in the human body, produced by folding of two alpha1(I) polypeptides and one alpha2(I) polypeptide into the triple helix. A conserved stem-loop structure is found in the 5' untranslated region of collagen mRNAs, encompassing the translation start codon. We cloned La ribonucleoprotein domain family member 6 (LARP6) as the protein that binds the collagen 5' stem-loop in a sequence-specific manner. LARP6 has a distinctive bipartite RNA binding domain not found in other members of the La superfamily. LARP6 interacts with the two single-stranded regions of the 5' stem-loop. The K(d) for binding of LARP6 to the 5' stem-loop is 1.4 nM. LARP6 binds the 5' stem-loop in both the nucleus and the cytoplasm. In the cytoplasm, LARP6 does not associate with polysomes; however, overexpression of LARP6 blocks ribosomal loading on collagen mRNAs. Knocking down LARP6 by small interfering RNA also decreased polysomal loading of collagen mRNAs, suggesting that it regulates translation. Collagen protein is synthesized at discrete regions of the endoplasmic reticulum. Using collagen-GFP (green fluorescent protein) reporter protein, we could reproduce this focal pattern of synthesis, but only when the reporter was encoded by mRNA with the 5' stem-loop and in the presence of LARP6. When the reporter was encoded by mRNA without the 5' stem-loop, or in the absence of LARP6, it accumulated diffusely throughout the endoplasmic reticulum. This indicates that LARP6 activity is needed for focal synthesis of collagen polypeptides. We postulate that the LARP6-dependent mechanism increases local concentration of collagen polypeptides for more efficient folding of the collagen heterotrimer.
Cai
Le
L
Department of Biomedical Sciences, College of Medicine, Tallahassee, FL 32306, USA.
Fritz
Dillon
D
Stefanovic
Lela
L
Stefanovic
Branko
B
D001324:Autoantigens / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000096:biosynthesis* / Q000235:genetics*; D017124:Conserved Sequence; D004247:DNA / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D000097585:SS-B Antigen
D001324:Autoantigens; D024042:Collagen Type I; D004247:DNA; D017931:DNA Primers; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D000097585:SS-B Antigen