Inhibition of hsp701 and hsp703 expression disrupts preimplantation embryogenesis and heightens embryo sensitivity to arsenic

Abstract

Mouse 70-kDa heat shock proteins Hsp70-1 and Hsp70-3 (Hsp70-1/3) are stress-inducible protein chaperones thought to protect embryonic cells and tissues from the effects of a wide range of environmental exposures. Hsp70-1/3 are expressed constitutively, and at times are stress-inducible during various stages of preimplantation embryogenesis. In order to elucidate the functions of constitutive and stress-inducible Hsp70 expression in mouse preimplantation embryos, the consequences of inhibiting expression with antisense oligonucleotides complementary to the mRNAs of hsp70-1 and hsp70-3 (A070-1/3) were evaluated. Transfection of preimplantation embryos (four-cell stage) with 2.5 microM A070-1/3 had no effect on in vitro blastocoel formation. However, transfection with 5 or 10 microM A070-1/3 reduced in vitro blastocyst development to 30% and 0%, respectively (approximately 90% control embryos developed to blastocyst). Thus constitutive expression of Hsp70-1/3 appears significant to preimplantation embryogenesis. Limiting expression of Hsp70-1/3 with 5 microM A070-1/3 also heightened embryo sensitivity to arsenic, resulting in less than 5% in vitro development to blastocyst in the presence of the subtoxic dose of 0.4 microM sodium arsenite. Whether the combined effect of A070-1/3 and arsenic is due to blocking inducible expression of the Hsp70s, or due to further reducing the amount of constitutively expressed Hsp70s available to the embryo is not known at this time. However, these results clearly indicate that some minimal amount of Hsp70-1 and/or Hsp70-3 is required for preimplantation embryogenesis, and that increasing the demand for Hsp70s by arsenic exposure heightens this requirement.

Heat shock proteins Hsp70‐1 and Hsp70‐3 Are necessary and sufficient to prevent arsenite‐induced dysmorphology in mouse embryos*

“…Using antisense oligonucleotides to inhibit expression of the stress-inducible Hsp70 proteins, the embryos were much more sensitive to arsenite than were conceptuses injected with a variety of control oligonucleotides. These results agree well with earlier results using the same oligonucleotides and preimplantation embryos exposed to arsenic (Dix et al, 1998). We postulate that the exposure of neuroepithelial cells to antisense oligonucleotide-lipid micelles within the amniotic cavity is suf®cient to affect gene expression and alter neural tube closure.…”

“…In vertebrates, Hsp70s appear to play a major role in thermotolerance in a variety of cell types (Anderson et al, 1989;Angelidis et al, 1991;Li et al, 1991) as well as in mouse oocytes (Hendrey and Kola, 1991). In cultured mouse preimplantation embryos, inhibition of Hsp70-1 and Hsp70-3 expression heightened embryo sensitivity to arsenic (Dix et al, 1998). Based upon this work with cells in culture, and the induction of Hsp70s in mammalian embryos, one of the obvious potential factors playing a role in embryonic thermotolerance is the stress-inducible Hsp70s.…”

Distinct Gene Expression Patterns of Two Heat Shock Protein 70 Members During Development, Diapause, and Temperature Stress in the Freshwater Crustacean Daphnia magna

“…The proteins of the different members of the Hsp70 family have been suggested to be involved in certain housekeeping mechanisms of embryogenesis, for instance, by moderating cell cycle, preventing apoptosis, and protecting embryos against environmental stress (Angelier et al, 1996;Dix et al, 1998;Zhu et al, 2013;Radons, 2016). This mirrors our results, as already from "stage I" we see the continuous presence of the protein encoded by the Dmhsp70-B gene.…”

Subcellular localization of presenilins during mouse preimplantation development

“…Antisense oligonucleotides have generated wide interest because of their potential to inhibit expression of specific genes in cells. Many studies have shown the roles of the important genes in the early development using antisense oligonucleotides (32,33). Our results suggest that both PS1 and 2 might be required coordinately for the cell division during the preimplantation development.…”