Satellite DNA binding and cellular localisation of RNA helicase P68.
We purified a 68-kDa protein from the mouse nuclear matrix using ion exchange and affinity chromatography. Column fractions were tested for specific binding to mouse minor satellite DNA using a gel mobility shift assay. The protein was identified by mass spectrometry as RNA helicase P68. In fixed cells, P68 was found to shuttle in and out of SC35 domains, forming fibres and granules in a cell-cycle dependent manner. Analysis of the P68 sequence revealed a short potential coiled-coil domain that might be involved in the formation of P68 fibres. Contacts between centromeres and P68 granules were observed during all phases of the cycle but they were most prominent in mitosis. At this stage, P68 was found in both the centromeric regions and the connections between chromosomes. Direct interaction of P68/DEAD box RNA helicase with satellite DNAs in vitro has not been demonstrated for any other members of the RNA helicase family.
Enukashvily
Natella
N
Cell Cultures Department, Institute of Cytology, Tikhoretsky, 4, St Petersburg, 194064, Russia. natella@mail.ctyspb.rssi.ru
Donev
Rossen
R
Sheer
Denise
D
Podgornaya
Olga
O
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002453:Cell Cycle / Q000502:physiology*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism; D002503:Centromere / Q000378:metabolism; D053487:DEAD-box RNA Helicases; D004276:DNA, Satellite / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D034882:Lamins / Q000378:metabolism; D008099:Liver / Q000737:chemistry; D051379:Mice; D008938:Mitosis / Q000502:physiology; D008969:Molecular Sequence Data; D015530:Nuclear Matrix / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000235:genetics; D020365:RNA Helicases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D004276:DNA, Satellite; D034882:Lamins; D011494:Protein Kinases; C506749:Ddx5 protein, human; C506751:Ddx5 protein, mouse; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases