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Report

Teaser, summary, work performed and final results

Periodic Reporting for period 1 - TFs-HSCs (The identification of the Combinatorial Transcription Factor network exerting different roles in Hematopoietic Stem Cells.)

Teaser

Summary

Transcription factors (TFs) are the proteins which bind DNA and have many roles. Among others, they regulate transcription and control various important biological processes. Our genome has ~22,000 genes, with ~1,400 encoding TFs. In most of the cases, TFs exert their functions by forming networks, termed as transcriptional-regulatory-networks (TRNs). However most of the studies have been focused on some TFs and their functions in biological processes like generation of the blood cells from the hematopoietic stem cells (HSCs), generation/expansion of HSCs and differentiation of embryonic stem cells (ESCs). ESCs are pluripotent cells that can generate all cell lineages, whereas HSCs can give rise to the different types of blood cells. The ramifications of inappropriate regulation of gene expression controlled by the TFs in such cell types can be in various levels; from no effect on the cells up to inappropriate behaviour of the cells leading to diseases and cancer. Hence, it is important to uncover all the uncharacterized TFs as well as the TRN involved in such processes (like maintenance and differentiation of HSCs) and decipher their role.

Work performed

A sgRNA library was designed based on the recent technological advances and updated predictions of the existing list of the TFs. That library would be subsequently used to apply the state-of-the-art CRISPR/Cas9 technology. At the same time, we focused on two TFs in order to evaluate the effect of their depletion in the regulation of transcription and in the biological processes and to evaluate the rules/criteria of the design of the sgRNAs. With ChIP-seq, we have identified the genome-wide localization of those TFs, the preferred regulatory elements that they target as well as their target genes. Furthermore, by combining ChIP-seq with RNA-seq, we have identified that those TFs (as a network) target genes essential for important biological processes. Thus, we have revealed that the two TFs maintain cells in an optimal configuration by controlling and regulating genes important for general functions, indicating the significance of TFs (and potentially TRNs) in governing important biological processes. Part of the results, will be presented in an upcoming conference from where I have received an invitation.

Final results

The common and recommended treatment for acute myeloid leukemia (AML) patients consists of chemotherapy followed by transplantation of HSCs. However, that results in low numbers of successfully transplanted HSCs and various complications. Thus, understanding the underlying mechanisms of the TFs generating/proliferating HSCs, generating the blood cells from HSCs and potentially generating HSCs from ESCs, will have important future applications in the clinic as well as socio-economic impact (e.g. transplantation of HSCs to AML patients).
For academia, the results will provide a list of of uncharacterized TFs and an insight into their function and their role in important biological processes, like generation of all cell lineages or generation of blood cells from ESCs or HSCs. The sgRNA library would be a valuable recourse for other scientists who would like to interrogate those TFs. For industry, the results could lead to new screening approaches and potentially developing new drugs, compounds and/or inhibitors.