Figure 1.
General organization of the hsp70 cluster in camel and human.
A – restriction maps of two overlapping recombinant phages, C3 and N10, used in the analysis (H – HindIII, X – XhoI, R – EcoRI, B – BamHI). B – general structure of the C. dromedarius HspA1 cluster. C – general structure of H. sapiens HSPA1 cluster provided for comparison. The length of the intergenic region between hspA1A and hspA1B genes is given in bps.
Figure 2.
Southern blot of camel genomic DNA with PCR-probe to hspA1A/B genes.
1 – BamHI, 2 – XbaI, 3 – XbaI/BamHI, 4 – EcoRI. M – fragment length markers.
Table 1.
Identity of camel hspA1 genes with orthologues from other organisms.
Table 2.
The structure of Hsp70 translation start and surrounding sequences in various organisms.
Table 3.
List of primers used in the experiments.
Figure 3.
Comparison of 5′-regulatory regions of hspA1 genes from camel and other mammals.
(A) Identification of conserved motifs in the region between hsp1L and hsp1A (designated LA) and upstream of hspA1B (designated B) from Camelus dromedarius (C), Bos taurus (B), Sus scrofa (S), Equus caballus (E), Homo sapiens (H), Mus musculus (M), Pteropus vampyrus (P), Tursiops truncatus (T), and Canis familiaris (D). Transcription start sites are indicated by arrows, and ATG codons – by triangles. Intron sequences of hsp1L genes (located 16 bp upstream from the ATG codon, as indicated by a vertical dotted line) were removed to reduce sequence heterogeneity. Motifs are numbered in the order of identification by MEME, and numbers at the bottom indicate approximate base pairs in the alignment. (B) Matches between selected motifs from panel A and binding sites of known transcription factors in the TRANSFAC database identified by TOMTOM. Shown are the logos with the corresponding p- and q-values for each TF. The remaining motifs do not yield any matches to binding sites of known TFs.
Figure 4.
A – RT-PCR with total RNA from camel's blood.
B – RT-PCR with total RNA from camel's heart muscle. 1 – primers CamORF1/2 to hspA1A/B genes and grp78, 2 – primers PT-1A and RT-2A to hspA1A/B genes, and 3 – primers RT-1L and RT-2L to hspA1L (see Table 3). RT – negative control of RT-PCR, samples without reverse transcriptase.
Figure 5.
Hsp70 family proteins isolated from camel heart.
The identity of proteins was determined by trypsin fingerprinting and microsequencing. Lane 2 – molecular weight marker.