Developmental causes of syndromic Thrombocytopenia and co-morbidities

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Congenital hematopoietic defects have been linked to a variety of co-morbidities affecting cardiovascular lineages, the kidneys, and limbs; curiously, their progenitor cells in the developing vertebrate embryo share a common embryonic origin in the lateral plate mesoderm (LPM). Understanding the developmental connection between hematopoietic lineage determination and other organ precursors forming from in part joint progenitor cells promises to shed light on the mechanisms of syndromic hematopoietic disorders. Thrombocytopenia Absent Radius (TAR) syndrome is a rare, congenital malformation that manifests as blood platelet defects and absent radius bones, concurrent with heart and kidney anomalies. TAR syndrome associates with chromosome 1q21.1 microdeletions that remove at least 20 genes including the exon junction complex (EJC) gene RBM8A/Y14 involved in splicing and mRNA degradation. Combined 1q21.1 microdeletion with compound inheritance of hypomorphic RBM8A alleles has found in a cohort of TAR patients, but how perturbed RBM8A/Y14 as general ubiquitous mRNA-processing factor causes the selective TAR phenotype remains to be uncovered. The syndrome’s phenotypes provide a paradigm for syndromic hematopoietic defects with complex co-morbidities which suggest that hypomorphic RBM8A perturbation predominantly impairs the lateral plate LPM when forming the progenitors for blood, limb, heart, and kidney.
Using zebrafish as a model system and our leading CRISPR-Cas9-based protocols, we have successfully established a first mutant allelic series for rbm8a. In live in toto imaging of developing rbm8a mutants in transgenic zebrafish reporter strains, we detected migration defects of the hematopoietic progenitor-harboring LPM stripes, indicating an early defect in LPM patterning. Our data define the first embryonic rbm8a phenotypes in the LPM and hematopoietic progenitors using zebrafish as in vivo platform to identify the developmental cause of distinctive hematopoietic lineage perturbation in TAR syndrome. Mechanistically, from our transcriptome analysis of rbm8a-mutant zebrafish embryos, we found that planar cell polarity (PCP) signaling components are deregulated, providing a key candidate pathway for perturbed hematopoietic progenitor migration in rbm8a mutants. We hypothesize that PCP/non-canonical WNT signaling results in a selective, pathological phenotype in the LPM. We will identify the mechanism as to how select mRNAs, in particular components of PCP signaling, are perturbed in rbm8a mutants, resulting in higher sensitivity to migration defects using genetic interaction studies and enhanced crosslinking immunoprecipitation (eCLIP). Our results will connect the control of early PCP/non-canonical WNT-based LPM migration to consequences in hematopoietic progenitor formation and establish a proxy to better understand the causes and connections between hematological and related phenotypes of syndromic birth defects.

Grant Number: 5R01DK129350-03
NIH Institute/Center: NIH
Principal Investigator: Alexa Burger