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Published on: January 19, 2021
Rapid DNA Synthesis During Early Drosophila Embryogenesis Is Sensitive to Maternal Humpty Dumpty Protein Function
Shera Lesly1, Jennifer L Bandura2, Brian R Calvi3
1Department of Biology, Indiana University, Bloomington, Indiana 47408.
Abstract:
Problems with DNA replication cause cancer and developmental malformations. It is not fully understood how DNA replication is coordinated with development and perturbed in disease. We had previously identified the Drosophila gene humpty dumpty (hd), and showed that null alleles cause incomplete DNA replication, tissue undergrowth, and lethality. Animals homozygous for the missense allele, hd , were viable, but adult females had impaired amplification of eggshell protein genes in the ovary, resulting in the maternal effects of thin eggshells and embryonic lethality. Here, we show that expression of an hd transgene in somatic cells of the ovary rescues amplification and eggshell synthesis but not embryo viability. The germline of these mothers remain mutant for the hd allele, resulting in reduced maternal Hd protein and embryonic arrest during mitosis of the first few S/M nuclear cleavage cycles with chromosome instability and chromosome bridges. Epistasis analysis of hd with the rereplication mutation plutonium indicates that the chromosome bridges of hd embryos are the result of a failed attempt to segregate incompletely replicated sister chromatids. This study reveals that maternally encoded Humpty dumpty protein is essential for DNA replication and genome integrity during the little-understood embryonic S/M cycles. Moreover, the two hd maternal-effect phenotypes suggest that ovarian gene amplification and embryonic cleavage are two time periods in development that are particularly sensitive to mild deficits in DNA replication function. This last observation has broader relevance for interpreting why mild mutations in the human ortholog of humpty dumpty and other DNA replication genes cause tissue-specific malformations of microcephalic dwarfisms.
Insights
Maternally inherited Humpty dumpty (hd) protein is crucial for embryonic DNA replication and genome stability. Mild deficits in hd function during ovarian gene amplification and early embryonic development can lead to developmental defects.
Area of Science:
- Developmental Biology
- Genetics
- Molecular Biology
Background:
- DNA replication errors are linked to cancer and developmental disorders.
- The precise coordination of DNA replication with development and its disruption in disease remain incompletely understood.
- The Drosophila gene humpty dumpty (hd) was previously identified, with null alleles causing incomplete DNA replication and lethality.
Purpose of the Study:
- To investigate the role of the humpty dumpty (hd) gene in DNA replication, development, and disease.
- To elucidate the function of maternally encoded Humpty dumpty protein during early embryonic development.
- To understand the basis of embryonic lethality and chromosome instability in hd mutants.
Main Methods:
- Genetic analysis in Drosophila melanogaster.
- Transgenic rescue experiments in somatic cells of the ovary.
- Epistasis analysis with the rereplication mutation plutonium.
- Microscopy to observe chromosome segregation and embryonic development.
Main Results:
- A missense allele of hd (hd) in females caused impaired ovarian gene amplification, thin eggshells, and embryonic lethality.
- Ovarian somatic cell expression of an hd transgene rescued amplification but not embryo viability.
- Mutant germline resulted in embryonic arrest during early cleavage cycles, exhibiting chromosome instability and bridges.
- Epistasis analysis revealed chromosome bridges result from failed segregation of incompletely replicated sister chromatids.
Conclusions:
- Maternally provided Humpty dumpty protein is essential for embryonic DNA replication and genome integrity during early S/M cycles.
- Ovarian gene amplification and embryonic cleavage are sensitive developmental periods to DNA replication deficits.
- Mild mutations in human orthologs of hd and other DNA replication genes may contribute to microcephalic dwarfisms and other malformations.
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