tailieunhanh - Báo cáo khoa học: Dissociation of DNA polymerase a-primase complex during meiosis in Coprinus cinereus

Previously, the activity of DNA polymeraseawas found in the meiotic prophase I including non-S phase stages, in the basidiomycetes,Coprinus cinereus. To study DNA poly-merasea during meiosis, we cloned cDNAs for the C. cinereusDNApolymeraseacatalytic subunit (p140) and C. cinereusprimase small subunit (p48). Northern analysis indicated that both p140 and p48 are expressed not only at S phase but also during the leptotene/zygotene stages of meiotic prophase I. | Eur. J. Biochem. 270 2137-2146 2003 FEBS 2003 doi Dissociation of DNA polymerase a-primase complex during meiosis in Coprinus cinereus Satoshi Namekawa Fumika Hamada Tomoyuki Sawadot Satomi Ishii Takayuki Narai Takashi Ishizaki Takashi Ohuchi Takao Arai and Kengo Sakaguchi Department of Applied Biological Science Faculty of Science and Technology Tokyo University of Science Japan Previously the activity of DNA polymerase a was found in the meiotic prophase I including non-S phase stages in the basidiomycetes Coprinus cinereus. To study DNA polymerase a during meiosis we cloned cDNAs for the C. cinereus DNA polymerase a catalytic subunit p140 and C. cinereus primase small subunit p48 . Northern analysis indicated that both p140 and p48 are expressed not only at S phase but also during the leptotene zygotene stages of meiotic prophase I. In situ immuno-staining of cells at meiotic prophase I revealed a sub population of p48 that does not colocalize with p140 in nuclei. We also purified the pol a-primase complex from meiotic cells by column chromatography and characterized its biochemical properties. We found a subpopulation of primase that was separated from the pol a-primase complex by phosphocellulose column chromatography. Glycerol gradient density sedimentation results indicated that the amount of intact pol a-primase complex in crude extract is reduced and that a smaller complex appears upon meiotic development. These results suggest that the form of the DNA polymerase a-primase complex is altered during meiotic development. Keywords meiotic prophase I zygotene pachytene pol a catalytic subunit p140 primase small subunit p48 . The DNA polymerase a-pol a -primase complex plays an essential role in eukaryotic DNA replication and the structural and biochemical properties of pol a are conserved across a wide range of eukaryotes 1 2 . According to the current model replicative DNA synthesis initiates from a short stretch of RNA primer