Functional characterization of two flap endonuclease-1 homologues in rice.
Flap endonuclease-1 (FEN-1) is an important enzyme involved in DNA replication and repair. Previously, we isolated and characterized a complementary DNA (cDNA) from rice (Oryza sativa) encoding a protein which shows homology with the eukaryotic flap endonuclease-1 (FEN-1). In this report, we found that rice (O. sativa L. cv. Nipponbare) possessed two FEN-1 homologues designated as OsFEN-1a and OsFEN-1b. The OsFEN-1a and OsFEN-1b genes were mapped to chromosome 5 and 3, respectively. Both genes contained 17 exons and 16 introns. Alignment of OsFEN-1a protein with OsFEN-1b protein showed a high degree of sequence similarity, particularly around the N and I domains. Northern hybridization and in situ hybridization analysis demonstrated preferential expression of OsFEN-1a and OsFEN-1b in proliferating tissues such as the shoot apical meristem or young leaves. The levels of OsFEN-1a and OsFEN-1b expression were significantly reduced when cell proliferation was temporarily halted by the removal of sucrose from the growth medium. When the growth-halted cells began to regrow following the addition of sucrose to the medium, both OsFEN-1a and OsFEN-1b were again expressed at high level. These results suggested that OsFEN-1a and OsFEN-1b are required for cell proliferation. Functional complementation assay suggested that OsFEN-1a cDNA had the ability to complement Saccharomyces cerevisiae rad27 null mutant. On the other hand, OsFEN-1b cDNA could not complement the rad27 mutant. The roles of OsFEN-1a and OsFEN-1b in plant DNA replication and repair are discussed.
Kimura
Seisuke
S
Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Chiba, 278-8510, Noda, Japan.
Furukawa
Tomoyuki
T
Kasai
Nobuyuki
N
Mori
Yoko
Y
Kitamoto
Hiroko K
HK
Sugawara
Fumio
F
Hashimoto
Junji
J
Sakaguchi
Kengo
K
D000595:Amino Acid Sequence; D002455:Cell Division / Q000235:genetics; D002874:Chromosome Mapping; D032461:Chromosomes, Plant / Q000235:genetics; D003001:Cloning, Molecular; D005091:Exons; D045585:Flap Endonucleases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics; D005816:Genetic Complementation Test; D017403:In Situ Hybridization; D007438:Introns; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D012275:Oryza / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D010802:Phylogeny; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D007527:Isoenzymes; D010940:Plant Proteins; D045585:Flap Endonucleases