Isolation and characterization of a sea urchin hsp 70 gene segment (original) (raw)
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Studies on heat shock proteins in sea urchin development
Development, Growth and Differentiation, 1999
Work on stress proteins in sea urchin embryos carried out over the last 20 years is reviewed and the following major results are described. Entire sea urchin embryos, if subjected to a rise in temperature at any postblastular stage undergo a wave of heat shock protein (hsp) synthesis and survive. If subjected to the same rise between fertilization and blastula formation, they are not yet able to synthesize hsp and die. Four clones coding for the major hsp, hsp70, have been isolated and sequenced; evidence for the existence of a heat shock factor has been provided, and a mechanism for the developmental regulation of hsp synthesis discussed. Intraembryonic and intracellular hsp location has been described; and a mechanism for achievement of thermotolerance proposed. A chaperonine role for a constitutive mitochondrial hsp56 has been suggested, as well as a role for the constitutive hsp70 in cell division. Heat shock, if preceded by 12-O-tetradecanoylphorbol-12-acetate (TPA) treatment causes apoptosis.
Biological research, 2018
Heat stress proteins are implicated in stabilizing and refolding denatured proteins in vertebrates and invertebrates. Members of the Hsp70 gene family comprise the cognate heat shock protein (Hsc70) and inducible heat shock protein (Hsp70). However, the cDNA sequence and the expression of Hsp70 in the Antarctic sea urchin are unknown. We amplified and cloned a transcript sequence of 1991 bp from the Antarctic sea urchin Sterechinus neumayeri, experimentally exposed to heat stress (5 and 10 °C for 1, 24 and 48 h). RACE-PCR and qPCR were employed to determine Hsp70 gene expression, while western blot and ELISA methods were used to determine protein expression. The sequence obtained from S. neumayeri showed high identity with Hsp70 members. Several Hsp70 family features were identified in the deduced amino acid sequence and they indicate that the isolated Hsp70 is related to the cognate heat shock protein type. The corresponding 70 kDa protein, called Sn-Hsp70, was immune detected in ...
Molecular cloning and expression of two HSP70 genes in the prawn, Macrobrachium rosenbergii
Cell Stress & Chaperones, 2004
Two complementary deoxyribonucleic acid (cDNA) clones encoding 2 different 70-kDa heat shock proteins (HSPs) were isolated from the prawn Macrobrachium rosenbergii. The cDNA clones were 2448 and 2173 bp in length and contained 1950-and 1734-bp open reading frames (ORFs), respectively. The ORFs encoded 649-and 577-amino acid polypeptides, which were named Mar-HSC70 and Mar-HSP70, respectively, according to the sequence identities with other known HSC70s and HSP70s and based on their inducibility in response to heat shock stress (at 35ЊC). Genomic DNA sequence analysis revealed no introns in either gene. The major structural differences between the 2 proteins were a 60-amino acid segment and a 14-amino acid segment present in the N-terminal and C-terminal, respectively, of Mar-HSC70 that were not found in Mar-HSP70. Northern blotting and semiquantitative reverse transcription-polymerase chain reaction analyses indicated that the Mar-HSP70 gene was expressed under heat shock (35ЊC) stress in a non-tissue-specific manner. In contrast, Mar-HSC70 messenger ribonucleic acid was constitutively expressed in every tissue except muscle, and its expression in response to heat shock (at 35ЊC) changed only in muscle.
Cell Stress and Chaperones, 2011
The heat shock proteins (HSPs) are a family of proteins whose expression is enhanced in response to environmental stressors. The Apostichopus japonicus hsp90 and hsp26 genes were cloned using expressed sequence tag and rapid amplification of cDNA ends techniques. The full-length cDNA of Aphsp90 and Aphsp26 contains 3,458 and 1,688 nucleotides encoding 720 and 236 amino acids, respectively. Multiple alignments indicated that the deduced amino acid sequences of ApHsp90 and ApHsp26 shared a high level of identity with Hsp90 and small SHPs (sHSPs) sequences of zebrafish, ant, acorn worms, etc., and shared identical structural features with Hsp90 and sHSPs. The expression profiles of these two genes under heat treatment were investigated by real-time quantitative PCR. It was found that the messenger RNA (mRNA) transcripts of the two A. japonicus genes varied among different tissues under normal conditions and heat shock, and that the mRNA expression of the two genes was higher in the intestine compared to other tissues. Heat shock significantly elevated the expression of Aphsp90 and Aphsp26 mRNA in a temperature-and time-dependent manner. The results indicate that Aphsp90 and Aphsp26 played important roles in mediating the environmental stress in A. japonicus.
Identification and Characterization of a Constitutive HSP75 in Sea Urchin Embryos
Biochemical and Biophysical Research Communications, 1997
An antiserum against a hsp of the 70-kDa family was prepared, by means of a fusion protein, which was able to detect a constitutive 75-kDa hsc in the sea urchin P. lividus. This hsc was present both during oogenesis and at all developmental stages. A two-dimensional electrophoresis has revealed four isolectric forms of this 75-kDa hsc. The amino acid sequence of the fragment used to prepare the anti-hsp70 antibodies revealed a 43% identity with the corresponding part of sea urchin sperm receptor, and in mature eggs a brighter immunofluorescence was seen all around the cell cortex where the receptor for sea urchin sperm is localized. In oocytes the hsp75 was localized in the cytoplasms but not in the nuclei. In the embryos a higher hsp75 concentration was found in the portion facing the lumen of the cells which invaginate at gastrulation.
Molecular and Cellular Biology, 1985
A hybrid gene in which the expression of an Escherichia coli j-galactosidase gene was placed under the control of a Drosophila melanogaster 70,000-dalton heat shock protein (hsp70) gene promoter was constructed. Mutant derivatives of this hybrid gene which contained promoter sequences of different lengths were prepared, and their heat-induced expression was examined in D. melanogaster and COS-1 (African green monkey kidney) cells. Mutants with 5' nontranscribed sequences of at least 90 and up to 1,140 base pairs were expressed strongly in both cell types. Mutants with shorter 5' extensions (of at least 63 base pairs) were transcribed and translated efficiently in COS-1 but not at all in D. melanogaster cells. Thus, in contrast to the situation in COS-1 cells, the previously defined heat shock consensus sequence which is located between nucleotides 62 and 48 of the hsp70 gene 5' nontranscribed DNA segment is not sufficient for the expression of the D. melanogaster gene in homologous cells. A second consensus-like element 69 to 85 nucleotides upstream from the cap site is postulated to be also involved in the heat-induced expression of the hsp7O gene in D. melanogaster cells.
Studies of cloned sequences from four drosophila heat shock loci
Cell, 1979
DNA cloned from the D. melanogaster (Oregon R) heat shock loci at 63BC and 950 codes for the 83,000 and the 68,000 dalton heat shock proteins, respectively. Both coding sequences occur once per haploid genome. Sequences complementary to messenger RNA for the 70,000 dalton heat shock protein are represented five times, twice at 87A and three times at 87C. The copies at 87A differ characteristically from those at 87C in an interval of a few hundred bp near the 5' end of the messenger sequence, and the corresponding two classes of hsp 70 messenger RNA are found on polysomes after heat shock. Within this differential region, there is about 15% divergence between messenger sequences cloned from the two loci, while in the rest of the messenger region examined the homology is much closer although still imperfect. Unexpectedly, considerable homology is found between the sequence for the 88,000 dalton heat shock protein at g5D and the sequences for the 70,000 dalton protein at 87A and 87C, and between these sequences and a site in 87D. Messenger RNA molecules of 2.4,2.55 and 3.05 kb code for the 68,000, 70,000 and 83,000 dalton heat shock proteins and hybridize to apparently uninterrupted DNA sequences of 2.1, 2.25 and 2.6 kb, respectively.
Transcription of sea urchin histone gene after heat shock
Molecular Biology Reports
Mesenchyme blastula sea-urchin embryos were heat-shocked at 31 "C and pulse-labelled with [3H]uridine. The nuclear RNA was fractionated on glyoxal/agarose gels and each RNA fraction hybridized with cloned early blastula histone DNA. The results showed that after heat shock there is an accumulation of histone RNA molecules larger than the messenger and a decrease, with respect to the control, of 9-S histone RNA. Chasing of the heat shock RNA by incubation of the embryos with unlabelled uridine and cytidine restores the radioactivity, as well as the hybridization profiles, of control embryos. Furthermore saturation curves obtained by hybridizing histone DNA with labelled 9-S RNA, showed an absence of the histone messenger in the cytoplasm of heat-shocked embryos, whereas it is present in both control and chased embryos. These results are consistent with the hypothesis that the accumulated histone RNA in heat-shocked embryos is the precursor of mature messengers.