Molecular characterization of the heat-inducible LmSTI1 protein of Leishmania major.
We have recently isolated a cDNA encoding the Leishmania major homologue of the yeast stress-inducible protein STI1. Southern blot analyses indicate that this protein is encoded by a single copy gene in L. major and that this gene is highly conserved throughout the Leishmania genus. The STI1 gene is constitutively expressed in both L. major promastigotes and amastigotes however, STI1 transcript levels can be upregulated in promastigotes by a shift in culture temperature from 26 to 37 degrees C. Upregulation of transcript was detectable within 5' of heat shock and continued to increase for a further 8 h before returning to constitutive levels. In addition, biosynthetic incorporation of [35S]methionine followed by immunoprecipitation revealed an increase in the level of nascent STI1 protein synthesized when promastigote cultures were shifted from 26 to 37 degrees C. The L. major STI1 protein and the heat shock proteins Hsp83 and Hsp70 form a salt-sensitive complex in L. major promastigotes as evidenced by co-immunoprecipitation using an antiserum specific for L. major STI1. Furthermore, this complex can be reconstituted in vitro by adding recombinant STI1 containing an amino-terminal histidine tag to promastigote lysate and subsequent purification using metal chelate affinity chromatography.
Webb
J R
JR
Infectious Disease Research Institute, Seattle, WA 98104, USA.
Campos-Neto
A
A
Skeiky
Y A
YA
Reed
S G
SG
D000595:Amino Acid Sequence; D000818:Animals; D016054:DNA, Protozoan / Q000032:analysis; D018628:Gene Dosage; D005786:Gene Expression Regulation / Q000502:physiology*; D017125:Genes, Protozoan / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics*; D006358:Hot Temperature; D007891:Leishmania / Q000235:genetics; D018320:Leishmania major / Q000235:genetics*; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D016053:RNA, Protozoan / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D016054:DNA, Protozoan; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C490310:Hsp83 protein, protozoan; D015800:Protozoan Proteins; D012333:RNA, Messenger; D016053:RNA, Protozoan; D011993:Recombinant Fusion Proteins; C109320:stress-inducible protein 1, Leishmania major