Characterization of the cis-acting element directing perinuclear localization of the metallothionein-1 mRNA.
Subcellular localization of mRNAs is a key mechanism for the synthesis of proteins close to their site of function. The mRNA encoding MT-1 (metallothionein-1) is localized in the perinuclear cytoplasm, where it is associated with cytoskeletal-bound polysomes. This localization relies on sequences present in the 3'-UTR (3'-untranslated region). The present study aims to characterize the cis-acting localization element(s) within the 3'-UTR. Using transfected cells expressing tagged MT-1 differing in their 3'-UTRs (deleted or mutated), the section(s) of this region required for directing MT-1 transcripts to the perinuclear cytoplasm has been investigated. Different 3'-UTRs were also used in UV cross-linking experiments that highlighted two distinct regions (nt 26-30 and 66-76) necessary for the binding of a protein of approx. 50 kDa, presumably involved in the mRNA targeting. The poor sequence homology between the MT-1 3'-UTR of various species, together with the bipartite nature of the required cis-element, indicates the involvement of a particular structure in the localization signal. The secondary structure of the MT-1 3'-UTR was investigated using enzymic and chemical probing. Current structural analysis of mutant 3'-UTRs will allow the critical structural features of the MT-1 mRNA perinuclear localization signal to be defined.
Chabanon
H
H
Institute of Cellular and Molecular Biosciences, University of Newcastle, Newcastle upon Tyne NE1 7RU, UK.
Nury
D
D
Mickleburgh
I
I
Burtle
B
B
Hesketh
J
J
D020413:3' Untranslated Regions; D000199:Actins / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D006801:Humans; D008668:Metallothionein / Q000096:biosynthesis* / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D011132:Polyribosomes / Q000737:chemistry; D017433:Protein Structure, Secondary; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D020218:Response Elements; D012689:Sequence Homology, Nucleic Acid; D014466:Ultraviolet Rays
D020413:3' Untranslated Regions; D000199:Actins; D003432:Cross-Linking Reagents; D012333:RNA, Messenger; D008668:Metallothionein