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Nuclear envelope associated protein that binds telomeric DNAs
Author(s) -
Podgornaya O.I.,
Bugaeva E.A.,
Voronin A.P.,
Gilson Eric,
Mitchell A.R.
Publication year - 2000
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/1098-2795(200009)57:1<16::aid-mrd4>3.0.co;2-8
Subject(s) - biology , microbiology and biotechnology , nuclear protein , telomere , chromatin , immunogold labelling , spermiogenesis , somatic cell , immunoprecipitation , cell nucleus , nucleus , dna , genetics , antibody , gene , transcription factor
Rana temporaria oocytes at the 6th diplotene stage of maturation contain a special structure, the karyosphere capsule, with chromosomes covered and detached from the nuclear envelope (NE), though at the previous stage the telomeres were attached to the membrane, as characteristic of germ cells. The DNA–protein complexes from band shift assays with proteins extracted from oocyte NEs and telomeric DNA fragment (T 2 G 4 ) 130 were isolated and injected into a guinea pig. In the present paper the only protein of 70 kDa recognized by antibody (AB) in the NE is named the Membrane Telomere Binding Protein (MTBP). Western blots with guinea pig AB and AB against telobox peptide from TRF2 show that protein of 60 kDa (probably TRF1) belongs to the chromatin, but MTBP (TRF2 according to immunoprecipitation) belongs to the NE. In the somatic cell nuclei both proteins are present and recognized by AB against telobox peptide, but AB raised recognize only MTBP/TRF2 due to the epitope different from telobox. Combined in situ hybridization with the vertebrate telomeric DNA sequences (T 2 AG 3 ) 135 and immunocytochemistry with the MTBP AB showed them to be colocalized within the mouse nucleus. As it was shown by immunofluorescense of NE spread, MTBP is organized in a distinct pattern that looks like a network made of double‐dots. Electron microscope immunogold staining with both ABs showed that the protein is localized on the outer surface of the oocyte NE within cup‐like structures attached to the membrane. This is the first clear evidence of a protein, which could be responsible for the attachment of telomeres to the nuclear membrane. Mol. Reprod. Dev. 57:16–25, 2000. © 2000 Wiley‐Liss, Inc.