Rewiring of RNA Polymerase II in a Novel Taspase1- Mediated Marrow Failure.

About This Grant

(PLEASE KEEP IN WORD, DO NOT PDF) Inherited Bone Marrow Failure Syndromes (IBMFS) constitute a rare set of disorders leading to an inability to generate functional circulating blood cells leading to morbidity and mortality. Many of these disorders are accompanied by pleiotropic developmental disorders. To date, we still lack definitive cures beyond highly toxic hematopoietic stem cell transplant. Decades of prior work has advanced our knowledge on these diseases and the fundamental roles that mutated causative genes play in normal hematopoiesis. However, nearly half of patients with IBMFS lack a clear genetic etiology, demonstrating the essential knowledge gap to understand genetic processes underlying marrow failure and devise new targeted therapies. Here, we present a novel, de novo, heterozygous mutation at Valine 343 to Methionine in Taspase1 (Tasp1V343M) in a child with an array of development anomalies and marrow failure. Tasp1V343M leads to failure of hematopoietic expansion and retention of early erythroid precursors in patient-derived iPSCs. Taspase1 is a highly conserved protease exerting effects on gene expression by proteolytic cleavage of target substrates. Molecular modeling demonstrates Tasp1V343M introduces intramolecular interactions leading to impaired flexibility of a domain allowing access to the proteolytic active site. Functionally, Tasp1V343M abrogates cleavage of TFIIA, a key member of the PreInitiation Complex (PIC) responsible for directing activation of RNA Polymerase II at promoters of mRNA. Furthermore, loss of TFIIA cleavage by Tasp1V343M leads to alterations of RNA Pol II promoter occupancy in iPSCs. These observations leave two unanswered questions of 1.) Does Tasp1V343M constitute a bone fide new marrow failure syndrome? and 2.) How does alteration of TFIIA reorchestrate transcriptomes in early development? Prior murine models of Taspase required homozygous deletion to manifest phenotypes implying Tasp1V343M is acting as a dominant negative. We will employ an array of genetic models including Tasp1V343M repair and non-cleavable TFIIA to demonstrate sufficiency of this mutation on hematopoiesis in vivo and delineate epistatic downstream mediators of Tasp1V343M marrow failure. Interestingly, noncleaved TFIIA has been postulated to direct RNA Pol II transcription in embryonic states independent of canonical PIC. We will investigate how uncleaved TFIIA alters RNA Pol II specificity genome wide, isolate determinants of target promoters and map these interactions across a model of erythropoiesis. This research will unlock a fundamental question on the diversity of mechanisms used to direct RNA Polymerase II in development and offer insight into a mechanism occurring early enough to impact pleiotropic tissue patterning and expand our knowledge of the pathophysiology of IBMFS.