A Telomere-Binding Protein (TRF2/MTBP) from Mouse Nuclear Matrix with Motives of an Intermediate Filament-Type Rod Domain

Abstract

In previous work, we identified a telomeric DNA-binding protein (termed telomere-membrane binding protein, MTBP) in the envelope of the frog oocyte nucleus and raised antibodies against it. Here we present immunological evidence which suggests strongly that MTBP is identical with the vertebrate telomeric DNA-binding protein TRF2 (telomere-repeat factor 2). MTBP/TRF2 possesses motif which resembles rod domain characteristic of intermediate filament (IF) proteins as shown by immunological cross-reactivity with characteristic antibodies, as well as amino acid sequence homology. Anti-MTBP antibodies recognised a protein of the same M, as TRF2 in extracts of mouse nuclei and nuclear matrix as shown by ion-exchange chromatography, gel shift assays, and Western blots. This mouse MTBP analogue forms more stable complexes with the vertebrate telomeric DNA fragment (T(2)AG(3))(135) than with the corresponding fragment from Tetrahymena (T(2)G(4))(130). Proteins in each of these complexes are recognised by anti-MTBP antibody. In situ hybridization with the vertebrate telomeric DNA sequence (T(2)AG(3))(135) and immunofluorescence with anti-MTBP antibody had shown earlier that these are co-localised in the nucleus of mouse cells, and here MTBP is shown to be associated with the residual membrane of hepatocyte nuclei using Western blotting and immunofluorescence. Some immunofluorescence signal from MTBP is localized at chromosome extremities on metaphase plates from mouse cell culture, but the main signal is seen in patches scattered around the chromosomes which were identified as remnants of the nuclear envelope by double labelling with antibodies against lamin B. These observations suggest that MTBP/TRF2 is a good candidate for the attachment of telomeres to the nuclear envelope in somatic cells.

Telomere-Binding TRF2/MTBP Localization during Mouse Spermatogenesis and Cell Cycle of the Mouse Cells L929

“…The TRF2/MTBP localizes at the ends of metaphase chromosomes. 18,37 MTBP antibodies (AB) revealed the same picture as TRF2 antibodies (Fig. 1, I 18 ).…”

Telomere-Binding TRF2/MTBP Localization during Mouse Spermatogenesis and Cell Cycle of the Mouse Cells L929

“…The TRF2/MTBP localizes at the ends of metaphase chromosomes. 18,37 MTBP antibodies (AB) revealed the same picture as TRF2 antibodies (Fig. 1, I 18 ).…”

Telomere and TRF2/MTBP Localization in Respect to Satellite DNA During the Cell Cycle of Mouse Cell Line L929

“…50,51 The TRF2 are located at the ends of metaphase chromosomes. 4,11 Part of the TRF2/MTBP antibody signals are seen at the ends of the chromosomes (Fig. 3 H,I) like those observed with antibody against TRF2.…”

Nucleolus-like body of mouse oocytes contains lamin A and B and TRF2 but not actin and topo II

“…TRF2 locations in somatic [ 44 ] and spermatogenic [ 64 , 65 ] cells have been determined. It has been suggested that TRF2 possesses intimate connections with the nuclear envelope [ 66 ].…”

“…In the diplotene stage, TRF1 remains connected with telomeres in chromatin, whereas TRF2/MTBP does not [ 43 ]. Its membrane location has been established by biochemical and morphological observations in mouse cells as well [ 44 , 64 ]. In mouse oocytes, TRF2 at stages 1 and 2 resides in SC35 domains (Fig.…”

Telomere-Binding TRF2/MTBP Localization during Mouse Spermatogenesis and Cell Cycle of the Mouse Cells L929

“…The TRF2/MTBP localizes at the ends of metaphase chromosomes. 18,37 MTBP antibodies (AB) revealed the same picture as TRF2 antibodies (Fig. 1, I 18 ).…”

Telomere and TRF2/MTBP Localization in Respect to Satellite DNA During the Cell Cycle of Mouse Cell Line L929

“…50,51 The TRF2 are located at the ends of metaphase chromosomes. 4,11 Part of the TRF2/MTBP antibody signals are seen at the ends of the chromosomes (Fig. 3 H,I) like those observed with antibody against TRF2.…”

Nucleolus-like body of mouse oocytes contains lamin A and B and TRF2 but not actin and topo II

“…TRF2 locations in somatic [ 44 ] and spermatogenic [ 64 , 65 ] cells have been determined. It has been suggested that TRF2 possesses intimate connections with the nuclear envelope [ 66 ].…”

“…In the diplotene stage, TRF1 remains connected with telomeres in chromatin, whereas TRF2/MTBP does not [ 43 ]. Its membrane location has been established by biochemical and morphological observations in mouse cells as well [ 44 , 64 ]. In mouse oocytes, TRF2 at stages 1 and 2 resides in SC35 domains (Fig.…”

Telomere-Binding TRF2/MTBP Localization during Mouse Spermatogenesis and Cell Cycle of the Mouse Cells L929

“…The TRF2/MTBP localizes at the ends of metaphase chromosomes. 18,37 MTBP antibodies (AB) revealed the same picture as TRF2 antibodies (Fig. 1, I 18 ).…”

Telomere and TRF2/MTBP Localization in Respect to Satellite DNA During the Cell Cycle of Mouse Cell Line L929

“…50,51 The TRF2 are located at the ends of metaphase chromosomes. 4,11 Part of the TRF2/MTBP antibody signals are seen at the ends of the chromosomes (Fig. 3 H,I) like those observed with antibody against TRF2.…”

Nucleolus-like body of mouse oocytes contains lamin A and B and TRF2 but not actin and topo II

“…TRF2 locations in somatic [ 44 ] and spermatogenic [ 64 , 65 ] cells have been determined. It has been suggested that TRF2 possesses intimate connections with the nuclear envelope [ 66 ].…”

“…In the diplotene stage, TRF1 remains connected with telomeres in chromatin, whereas TRF2/MTBP does not [ 43 ]. Its membrane location has been established by biochemical and morphological observations in mouse cells as well [ 44 , 64 ]. In mouse oocytes, TRF2 at stages 1 and 2 resides in SC35 domains (Fig.…”