Murine cDNAs coding for the centrosomal antigen centrosomin A.
Screening of an induced Ehrlich ascites cell-derived lambda gt11 cDNA library with an antibody (GP1), immunoreacting specifically with centrosomal antigen(s) of interphase and mitotic cells of different species, released a partial cDNA clone (lambda P10A) encoding the carboxy-terminal section of a centrosome-specific antigen. This specificity of the clone lambda P10A could be verified by lacZ-directed antigen expression from Escherichia coli Y1089 lysogenized with the recombinant phage lambda P10A and subsequent production of centrosome-specific antibodies by means of the recombinant antigen. Using the lambda P10A insert as a probe, two types of cDNA clones were identified in a lambda gt10 cDNA library by plaque-hybridization. The inserts of PN1 type clones were 1.2 kb (kilobases) and those of PN5 type clones were 2.2 kb in length. The DNA sequence of a PN1 type clone revealed its full-length cDNA nature. The open reading frame of PN1 encodes a rather hydrophilic and highly charged 34.5 x 10(3) Mr polypeptide comprising short but apparently significant strings of 100% sequence identity with the major nuclear lamina polypeptides lamins A/C and lamin B. Restriction enzyme mapping of PN1 and PN5 inserts, cross-hybridization experiments and comparison of overlapping DNA sequences indicate that the 1.2 kb and 2.2 kb cDNAs code for the same 34.5 x 10(3) Mr polypeptide, termed centrosomin A. Western blots of Ehrlich ascites cell proteins show a second, larger GP1 antigen (centrosomin B) whose cDNA has not been cloned. It remains to be investigated whether centrosomin B is encoded by a second mRNA or whether it reflects an oligomeric or a postranslationally modified form of centrosomin A.
Joswig
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Institute of Cell and Tumor Biology, German Cancer Research Center, Heidelberg, FRG.
Petzelt
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Werner
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D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000941:Antigens / Q000235:genetics*; D034961:Antigens, Nuclear; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D015342:DNA Probes; D004274:DNA, Recombinant; D015723:Gene Library; D005815:Genetic Code / Q000502:physiology; D051379:Mice; D015335:Molecular Probes; D008969:Molecular Sequence Data; D015388:Organelles / Q000276:immunology*; D011993:Recombinant Fusion Proteins
D000911:Antibodies, Monoclonal; D000941:Antigens; D034961:Antigens, Nuclear; D015342:DNA Probes; D004274:DNA, Recombinant; D015335:Molecular Probes; D011993:Recombinant Fusion Proteins; C069516:centrosomin A; D004247:DNA