A physical and functional constituent of telomerase anchor site.
Telomerase is a ribonucleoprotein reverse transcriptase responsible for the maintenance of one strand of the telomere terminal repeats. It consists minimally of a catalytic protein component (TERT) and an RNA subunit that provides the template. Compared with prototypical reverse transcriptases, telomerase is unique in possessing a DNA binding domain (anchor site) that is distinct from the catalytic site. Yeast TERT mutants bearing deletion or point mutations in an N-terminal domain (known as N-GQ) were found to be selectively impaired in extending primers that form short hybrids with telomerase RNA. The mutants also suffered a significant loss of repeat addition processivity but displayed an enhancement in nucleotide addition processivity. Furthermore, the mutants manifested altered primer utilization properties for oligonucleotides containing non-telomeric residues in the 5'-region. Cross-linking studies indicate that the N-GQ domain physically contacts the 5'-region of the DNA substrate in the context of a telomerase-telomere complex. Together, these results implicate the N-GQ domain of TERT as a physical and functional constituent of the telomerase anchor site. Coupled with previous genetic analysis, our data confirm that anchor site interaction is indeed important for telomerase function in vivo.
Lue
Neal F
NF
Department of Microbiology and Immunology, W. R. Hearst Microbiology Research Center, Weill Medical College of Cornell University, New York, New York 10021, USA. nflue@med.cornell.edu
D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D020134:Catalytic Domain; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D016812:Euplotes / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry; D017353:Gene Deletion; D066298:In Vitro Techniques; D015233:Models, Statistical; D008969:Molecular Sequence Data; D009154:Mutation; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D019098:Telomerase / Q000737:chemistry* / Q000378:metabolism; D016615:Telomere / Q000648:ultrastructure; D013768:Tetrahymena / Q000378:metabolism
D003432:Cross-Linking Reagents; D017931:DNA Primers; D005656:Fungal Proteins; D009841:Oligonucleotides; D012313:RNA; D004247:DNA; D019098:Telomerase