Interplay between primase and replication factor C in the hyperthermophilic archaeon Sulfolobus solfataricus.
The heterodimeric primase from the hyperthermophilic archaeon Sulfolobus solfataricus synthesizes long RNA and DNA products in vitro. How primer synthesis by primase is coupled to primer extension by DNA polymerase in this organism is unclear. Here we show that the small subunit of the clamp loader replication factor C (RFC) of S. solfataricus interacted with both the catalytic and non-catalytic subunits of the primase by yeast two-hybrid and co-immunoprecipitation assays. Further, the primase-RFC interaction was also identified in the cell extract of S. solfataricus. Deletion analysis indicated that the small subunit of RFC interacted strongly with the N-terminal domain of the catalytic subunit of the primase. RFC stimulated dinucleotide formation but decreased the amount of primers synthesized by the primase. The inhibition of primer synthesis is consistent with the observation that RFC reduced the affinity of the primase for DNA templates. On the other hand, primase stimulated the ATPase activity of RFC. These findings suggest that the primase-RFC interaction modulates the activities of both enzymes and therefore may be involved in the regulation of primer synthesis and the transfer of primers to DNA polymerase in Archaea.
Wu
Kangyun
K
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100080, China.
Lai
Xiaoqin
X
Guo
Xin
X
Hu
Jinchuan
J
Xiang
Xiaoyu
X
Huang
Li
L
D000251:Adenosine Triphosphatases / Q000378:metabolism; D019843:Archaeal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D019915:DNA Primase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D051818:Replication Protein C / Q000302:isolation & purification / Q000378:metabolism*; D048229:Sulfolobus solfataricus / Q000201:enzymology / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D019843:Archaeal Proteins; C034477:RNA primers; D011994:Recombinant Proteins; D012313:RNA; D004247:DNA; D019915:DNA Primase; D000251:Adenosine Triphosphatases; D051818:Replication Protein C