Molecular characterization and ultrastructural localization of Plasmodium falciparum Hsp 60.
Heat shock proteins (Hsp) are a group of highly conserved proteins which are widely represented phylogenetically. Genes for members of the Hsp 70, 90 and 60 families have been cloned from the human malaria parasite Plasmodium falciparum. In this study, we have cloned and expressed the P. falciparum Hsp 60 (PfHsp60) in E. coli. The sequence analysis identified a previously unknown intron of 257 bp beginning after the nucleotide 142 in the coding sequence. Antisera raised against the recombinant PfHsp60 was employed in immunoprecipitation studies with biosynthetically labeled parasite extracts to investigate regulation of expression of PfHsp60 at various temperatures. In contrast to the three to four fold accumulation of PfHsp60 transcripts in heat shocked parasites (37-40 degrees C), the expression of PfHsp60 was not induced in the blood stages of P. falciparum. On the other hand, the effect of heat induction on PfHsp70 was seen both at the level of specific mRNA and protein. In these studies we also observed co-immunoprecipitation of a number of other cellular proteins suggesting possible interaction with PfHsp60. Immunofluorescence analysis indicated the presence of PfHsp60 in the cytoplasm of all the various stages of the parasite. In addition, immunoelectron microscopic analysis distinctly localized PfHsp60 in the mitochondrion of P. falciparum. This study suggests that different mechanisms are involved in the regulation of expression of various members of the heat shock proteins in the parasite.
Das
A
A
Department of Molecular Microbiology and Immunology, Johns Hopkins School of Public Health, Baltimore, MD 21205-2079, USA.
Syin
C
C
Fujioka
H
H
Zheng
H
H
Goldman
N
N
Aikawa
M
M
Kumar
N
N
D000818:Animals; D001483:Base Sequence; D018834:Chaperonin 60 / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D017125:Genes, Protozoan; D006358:Hot Temperature; D006801:Humans; D007438:Introns; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000235:genetics* / Q000378:metabolism* / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D015800:Protozoan Proteins / Q000235:genetics
D018834:Chaperonin 60; D017931:DNA Primers; D016054:DNA, Protozoan; D015800:Protozoan Proteins