Protein kinase CK2 phosphorylates the high mobility group domain protein SSRP1, inducing the recognition of UV-damaged DNA.
The structure-specific recognition protein SSRP1 plays a role in transcription and replication in the chromatin context. Mediated by its C-terminal high mobility group (HMG) box domain, SSRP1 binds DNA non-sequence specifically but recognizes certain DNA structures. Using acetic acid urea polyacrylamide gel electrophoresis and mass spectrometry, we have examined the phosphorylation of maize SSRP1 by protein kinase CK2 alpha. The kinase phosphorylated several amino acid residues in the C-terminal part of the SSRP1 protein. Two phosphorylation sites were mapped in the very C-terminal region next to the HMG box domain, and about seven sites are localized within the acidic domain. Circular dichroism showed that the phosphorylation of the two C-terminal sites by CK2 alpha resulted in a structural change in the region of HMG box domain, because the negative peak of the CD spectrum at 222 nm was decreased by approximately 10%. In parallel, the phosphorylation induced the recognition of UV-damaged DNA, whereas the non-phosphorylated protein does not discriminate between UV-damaged DNA and control DNA. The affinity of CK2 alpha-phosphorylated SSRP1 for the DNA correlates with the degree of UV-induced DNA damage. Moreover, maize SSRP1 can restore the increased UV-sensitivity of a yeast strain lacking the NHP6A/B HMG domain proteins to levels of the control strain. Collectively, these findings indicate a role for SSRP1 in the UV response of eukaryotic cells.
Krohn
Nicholas M
NM
Institute of Life Sciences, Aalborg University, Sohngaardsholmsvej 49, DK-9000 Aalborg, Denmark.
Stemmer
Christian
C
Fojan
Peter
P
Grimm
Rudi
R
Grasser
Klaus D
KD
D019943:Amino Acid Substitution; D001483:Base Sequence; D047390:Casein Kinase II; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism* / Q000528:radiation effects*; D004249:DNA Damage; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006609:High Mobility Group Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D010766:Phosphorylation; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D035602:Transcriptional Elongation Factors; D014466:Ultraviolet Rays; D003313:Zea mays / Q000378:metabolism
D017931:DNA Primers; D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; C073537:SSRP1 protein, human; D035602:Transcriptional Elongation Factors; D004247:DNA; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases