Functional organization of hsp70 cluster in camel (Camelus dromedarius) and other mammals

© The Author(s), 2011. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS One 6 (2011): e27205, doi:10.1371/journal.pone.0027205.Heat shock protein 70 (Hsp70) is a molecular chaperone providing tolerance to heat and other challenges at the cellular and organismal levels. We sequenced a genomic cluster containing three hsp70 family genes linked with major histocompatibility complex (MHC) class III region from an extremely heat tolerant animal, camel (Camelus dromedarius). Two hsp70 family genes comprising the cluster contain heat shock elements (HSEs), while the third gene lacks HSEs and should not be induced by heat shock. Comparison of the camel hsp70 cluster with the corresponding regions from several mammalian species revealed similar organization of genes forming the cluster. Specifically, the two heat inducible hsp70 genes are arranged in tandem, while the third constitutively expressed hsp70 family member is present in inverted orientation. Comparison of regulatory regions of hsp70 genes from camel and other mammals demonstrates that transcription factor matches with highest significance are located in the highly conserved 250-bp upstream region and correspond to HSEs followed by NF-Y and Sp1 binding sites. The high degree of sequence conservation leaves little room for putative camel-specific regulatory elements. Surprisingly, RT-PCR and 5′/3′-RACE analysis demonstrated that all three hsp70 genes are expressed in camel's muscle and blood cells not only after heat shock, but under normal physiological conditions as well, and may account for tolerance of camel cells to extreme environmental conditions. A high degree of evolutionary conservation observed for the hsp70 cluster always linked with MHC locus in mammals suggests an important role of such organization for coordinated functioning of these vital genes.This work was supported by the Russian Foundation for Basic Research, project 09-04-00643 and 09-04-00660, project from ‘‘Genofond dynamics’’ program, and Grant of the Program of Molecular and Cellular Biology RAN to Dr. Evgen’ev; and by the Ministry of Education and Science of Russian Federation (State contract 14.740.11.0757 and Russia President Grant to young scientists MK-1418.2010.4. The research was supported by State Contract N16.552.11.7034 of Ministry of Education and Science

Heat Shock Proteins and Whole Body Adaptation to Extreme Environments

XVII, 218 p. 47 illus., 38 illus. in color.onlin

Organization and evolution of <it>hsp70 </it>clusters strikingly differ in two species of Stratiomyidae (Diptera) inhabiting thermally contrasting environments

<p>Abstract</p> <p>Background</p> <p>Previously, we described the heat shock response in dipteran species belonging to the family Stratiomyidae that develop in thermally and chemically contrasting habitats including highly aggressive ones. Although all species studied exhibit high constitutive levels of Hsp70 accompanied by exceptionally high thermotolerance, we also detected characteristic interspecies differences in heat shock protein (Hsp) expression and survival after severe heat shock. Here, we analyzed genomic libraries from two Stratiomyidae species from thermally and chemically contrasting habitats and determined the structure and organization of their <it>hsp70 </it>clusters.</p> <p>Results</p> <p>Although the genomes of both species contain similar numbers of <it>hsp70 </it>genes, the spatial distribution of <it>hsp70 </it>copies differs characteristically. In a population of the eurytopic species <it>Stratiomys singularior</it>, which exists in thermally variable and chemically aggressive (hypersaline) conditions, the <it>hsp70 </it>copies form a tight cluster with approximately equal intergenic distances. In contrast, in a population of the stenotopic <it>Oxycera pardalina </it>that dwells in a stable cold spring, we did not find <it>hsp70 </it>copies in tandem orientation. In this species, the distance between individual <it>hsp70 </it>copies in the genome is very large, if they are linked at all. In <it>O. pardalina </it>we detected the <it>hsp68 </it>gene located next to a <it>hsp70 </it>copy in tandem orientation. Although the <it>hsp70 </it>coding sequences of <it>S. singularior </it>are highly homogenized via conversion, the structure and general arrangement of the <it>hsp70 </it>clusters are highly polymorphic, including gross aberrations, various deletions in intergenic regions, and insertion of incomplete <it>Mariner </it>transposons in close vicinity to the 3'-UTRs.</p> <p>Conclusions</p> <p>The <it>hsp70 </it>gene families in <it>S. singularior </it>and <it>O. pardalina </it>evolved quite differently from one another. We demonstrated clear evidence of homogenizing gene conversion in the <it>S. singularior hsp70 </it>genes, which form tight clusters in this species. In the case of the other species, <it>O. pardalina</it>, we found no clear trace of concerted evolution for the dispersed <it>hsp70 </it>genes. Furthermore, in the latter species we detected <it>hsp70 </it>pseudogenes, representing a hallmark of the birth-and-death process.</p

Remarkable Site Specificity of Local Transposition Into the Hsp70 Promoter of Drosophila melanogaster

Heat-shock genes have numerous features that ought to predispose them to insertional mutagenesis via transposition. To elucidate the evolvability of heat-shock genes via transposition, we have exploited a local transposition technique and Drosophila melanogaster strains with EPgy2 insertions near the Hsp70 gene cluster at 87A7 to produce numerous novel EPgy2 insertions into these Hsp70 genes. More than 50% of 45 independent insertions were made into two adjacent nucleotides in the proximal promoter at positions −96 and −97, and no insertions were into a coding or 3′-flanking sequence. All inserted transposons were in inverse orientation to the starting transposon. The frequent insertion into nucleotides −96 and −97 is consistent with the DNase hypersensitivity, absence of nucleosomes, flanking GAGA-factor-binding sites, and nucleotide sequence of this region. These experimental insertions recapitulated many of the phenotypes of natural transposition into Hsp70: reduced mRNA expression, less Hsp70 protein, and decreased inducible thermotolerance. The results suggest that the distinctive features of heat-shock promoters, which underlie the massive and rapid expression of heat-shock genes upon heat shock, also are a source of evolutionary variation on which natural selection can act

Rodent Models of Audiogenic Epilepsy: Genetic Aspects, Advantages, Current Problems and Perspectives

Animal models of epilepsy are of great importance in epileptology. They are used to study the mechanisms of epileptogenesis, and search for new genes and regulatory pathways involved in the development of epilepsy as well as screening new antiepileptic drugs. Today, many methods of modeling epilepsy in animals are used, including electroconvulsive, pharmacological in intact animals, and genetic, with the predisposition for spontaneous or refractory epileptic seizures. Due to the simplicity of manipulation and universality, genetic models of audiogenic epilepsy in rodents stand out among this diversity. We tried to combine data on the genetics of audiogenic epilepsy in rodents, the relevance of various models of audiogenic epilepsy to certain epileptic syndromes in humans, and the advantages of using of rodent strains predisposed to audiogenic epilepsy in current epileptology

Activity of heat shock genes' promoters in thermally contrasting animal species.

Heat shock gene promoters represent a highly conserved and universal system for the rapid induction of transcription after various stressful stimuli. We chose pairs of mammalian and insect species that significantly differ in their thermoresistance and constitutive levels of Hsp70 to compare hsp promoter strength under normal conditions and after heat shock (HS). The first pair includes the HSPA1 gene promoter of camel (Camelus dromedarius) and humans. It was demonstrated that the camel HSPA1A and HSPA1L promoters function normally in vitro in human cell cultures and exceed the strength of orthologous human promoters under basal conditions. We used the same in vitro assay for Drosophila melanogaster Schneider-2 (S2) cells to compare the activity of the hsp70 and hsp83 promoters of the second species pair represented by Diptera, i.e., Stratiomys singularior and D. melanogaster, which dramatically differ in thermoresistance and the pattern of Hsp70 accumulation. Promoter strength was also monitored in vivo in D. melanogaster strains transformed with constructs containing the S. singularior hsp70 ORF driven either by its own promoter or an orthologous promoter from the D. melanogaster hsp70Aa gene. Analysis revealed low S. singularior hsp70 promoter activity in vitro and in vivo under basal conditions and after HS in comparison with the endogenous promoter in D. melanogaster cells, which correlates with the absence of canonical GAGA elements in the promoters of the former species. Indeed, the insertion of GAGA elements into the S. singularior hsp70 regulatory region resulted in a dramatic increase in promoter activity in vitro but only modestly enhanced the promoter strength in the larvae of the transformed strains. In contrast with hsp70 promoters, hsp83 promoters from both of the studied Diptera species demonstrated high conservation and universality

Characteristic Features of the Transcriptome in a Rat Strain with Audiogenic Epilepsy

Audiogenic epilepsy (AE), developing in rodent strains in response to sound, is widely used as the model of generalized convulsive epilepsy, while the molecular mechanisms determining AE are currently poorly understood. The brain region that is crucial for AE development isthe inferior and superior colliculi (IC, SC). We compared IC-SC gene expression profiles in rats with different AE susceptibility using transcriptome analysis.The transcriptomes were obtained from the IC-SC of Wistar rats (with no AE), Krushinsky-Molodkina (KM) strain rats (100% AE susceptible), and ”0” strain rats (with no AE) selected from F2 KM x Wistar hybrids for AE absence. KM gene expression displayed characteristic differences inboth of the strains that were not susceptible to AE. There was increased expression of a number of genes responsible for positive regulation of the MAPK signaling cascade, as well as of genes responsible for the production of interferon and several other cytokines. An increase in the expression levels of theTTR gene was found in KM rats, as well as significantly lower expression of the Msh3 gene (involved in post-replicative DNA repair systems). AE was also describedin the 101/HY mouse strain with a mutation in the locus controlling DNA repair. The DNA repair system defects could be the primary factor leading to the accumulation of mutations, which, in turn, promote AE. Keywords: udiogenic seizure, KM strain, transcriptome, TTR gene, Msh3 gene, DNA repai