We propose Drosophila melanogaster as animal model to understand the biological causes of human autosomal recessive primary microcephaly (MCPH). All genes causing primary microcephaly (PM) in humans have Drosophila orthologues. To date, the majority of the MCPH-associated genes encode protein that play important roles in different aspect of cell division process, from centrosome biology to kinetochore structure to MT dynamics. Our research project is aimed to clarify the mechanisms underlying impaired neurogenesis in PM by understanding how defects in functionally different proteins impact on the limitation of neural stem cell division capacity. We suggest 2 hypotheses to explain this reduction in neuron number. 1) The genetic defects associated with PM generate high level of DNA damage both/or by affecting the ability to repair spontaneous double strand breaks (DSBs) generated during development and/or by causing chromosome mis-segregation; this genomic instability will ultimately trigger apoptosis leading to production of fewer neurons. 2) Mutations in MCPH-associated genes alter the balance between proliferative and differentiative cell state and induce premature neuron differentiation.
To understand how these biological processes are involved in PM genesis we plan (i) to analyze for both DNA Damage and apoptosis Drosophila mutants of conserved PM genes; (ii) to examine the effect of mutations in additional Drosophila mitotic genes on DNA integrity and cell viability (iii) to study the nuclear architecture by cytological analysis of the distribution of heterochromatic regions.
We believe that our results will shed new light on the genetic and molecular basis of PM.
In the primary microcephaly field, patients and physicians face two difficulties: diagnostic uncertainty and lack of specific treatment. Diagnosis is long and expensive, and is complicated by the relatively lower frequency of the genetic defects compared to the non-genetic microcephaly forms, as well as by the phenotypic heterogeneity of the genetic forms. For the same reasons, it is very difficult to provide patients with accurate prognostic estimates. By increasing our knowledge of pathogenetic microcephaly mechanisms, our project has the potential to improve diagnostic and prognostic accuracy in at least two ways. First, it will provide information about the cellular processes that are involved in the pathology of microcephaly. This knowledge will shed light on unknown genetic factors that can significantly modify the penetrance and expressivity of phenotypes. Second, it will provide new candidate genes, which could help refining the interpretation of sequence variants identified in patients by next generation sequencing efforts. In this respect, it must be considered that many clinical cases of genetic microcephaly remain unresolved even after undergoing exome or whole genome sequencing.
The development of effective therapeutic strategies for neuro-developmental disorders characterized by extensive loss of neural progenitors or neurons is one of the hardest challenges of modern medicine. In consideration of the dramatic technical and biological difficulties posed by gene therapy and by stem-cells transplantation, the development of pharmacological strategies aimed to interfere with some of the common mechanisms leading to reduced brain mass represents an appealing possibility. We believe that the identification of critical pathogenetic mechanisms leading to microcephaly could further highlight potential targets for therapeutic intervention.
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