A novel hnRNP protein (HAP/SAF-B) enters a subset of hnRNP complexes and relocates in nuclear granules in response to heat shock.
A two-hybrid screening in yeast for proteins interacting with the human hnRNP A1, yielded a nuclear protein of 917 amino acids that we termed hnRNP A1 associated protein (HAP). HAP contains an RNA binding domain (RBD) flanked by a negatively charged domain and by an S/K-R/E-rich region. In in vitro pull-down assays, HAP interacts with hnRNP A1, through its S/K-R/E-rich region, and with several other hnRNPs. HAP was found to be identical to the previously described Scaffold Attachment Factor B (SAF-B) and to HET, a transcriptional regulator of the Heat Shock Protein 27 gene. We show that HAP is a bona fide hnRNP protein, since anti-HAP antibodies immunoprecipitate from HeLa cell nucleoplasm the complete set of hnRNP proteins. Unlike most hnRNP proteins, the subnuclear distribution of HAP is profoundly modified in heat-shocked HeLa cells. Heat-shock treatment at 42 degrees C causes a transcription-dependent recruitment of HAP to a few large nuclear granules that exactly coincide with sites of accumulation of Heat Shock Factor 1 (HSF1). The recruitment of HAP to the granules is temporally delayed with respect to HSF1 and persists for a longer time during recovery at 37 degrees C. The hnRNP complexes immunoprecipitated from nucleoplasm of heat-shocked cells with anti-HAP antibodies have an altered protein composition with respect to canonical complexes. Altogether our results suggest an involvement of HAP in the cellular response to heat shock, possibly at the RNA metabolism level.
Weighardt
F
F
Istituto di Genetica Biochimica ed Evoluzionistica, CNR Via Abbiategrasso 207, Italy.
Cobianchi
F
F
Cartegni
L
L
Chiodi
I
I
Villa
A
A
Riva
S
S
Biamonti
G
G
D000595:Amino Acid Sequence; D002453:Cell Cycle; D002467:Cell Nucleus / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D007700:Kinetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D005810:Multigene Family; D012261:Ribonucleoproteins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D013997:Time Factors; D014158:Transcription, Genetic
D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D012261:Ribonucleoproteins