Subcellular structure containing mRNA for beta subunit of mitochondrial H+-ATP synthase in rat hepatocytes is translationally active.
We have recently reported that the nuclear-encoded mRNA for the beta subunit of mitochondrial H+-ATP synthase (beta-mRNA) is localized in rounded, electron-dense clusters in the cytoplasm of rat hepatocytes. Clusters of beta-mRNA are often found in close proximity to mitochondria. These findings suggested a role for these structures in controlling the cytoplasmic expression and sorting of the encoded mitochondrial precursor. Here we have addressed the question of whether the structures containing beta-mRNA are translationally active. For this purpose a combination of high-resolution in situ hybridization and immunocytochemical procedures was used. Three different co-localization criteria showed that beta-mRNA-containing structures always revealed positive immunoreactive signals for mitochondrial H+-ATP synthase (F1-ATPase), ribosomal and hsc70 proteins. Furthermore, clusters show evidence in situ of developmental changes in the translational efficiency of the beta-mRNA. These findings suggest that structures containing beta-mRNA are translationally active irrespective of their cytoplasmic location. The immunocytochemical quantification of the cytoplasmic presentation of hsc70 in the hepatocyte reveals that approx. 86% of the protein has a dispersed distribution pattern. However, the remaining hsc70 is presented in clusters of which only half reveal positive hybridization for beta-mRNA. The interaction of hsc70 with the beta-F1-ATPase precursor protein is documented by the co-localization of F1-ATPase immunoreactive material within cytoplasmic clusters of hsc70 and by the co-immunoprecipitation of hsc70 with the beta-subunit precursor from liver post-mitochondrial supernatants. Taken together, these results suggest a role for hsc70 in the translation/sorting pathway of the mammalian precursor of the beta-F1-ATPase protein.
Ricart
J
J
Departamento de Biología Molecular, Centro de Biología Molecular 'Severo Ochoa', Universidad Autónoma de Madrid, Consejo Superior de Investigaciones Científicas, Universidad Autónoma de Madrid, 28049 Madrid, Spain.
Egea
G
G
Izquierdo
J M
JM
San Martín
C
C
Cuezva
J M
JM
D000818:Animals; D000831:Animals, Newborn; D018076:DNA, Complementary / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000032:analysis; D017403:In Situ Hybridization; D008099:Liver / Q000378:metabolism*; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism; D015388:Organelles / Q000378:metabolism*; D010750:Phosphoproteins / Q000032:analysis; D014176:Protein Biosynthesis; D017510:Protein Folding; D006180:Proton-Translocating ATPases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012269:Ribosomal Proteins / Q000032:analysis
D018076:DNA, Complementary; D018840:HSP70 Heat-Shock Proteins; D010750:Phosphoproteins; D012333:RNA, Messenger; D012269:Ribosomal Proteins; D006180:Proton-Translocating ATPases