COMPUTATIONAL MODELLING OF SEQUENCE FEATURES DRIVING TRANSCRIPTION FACTOR BINDING IN NEURONAL REPROGRAMMING
Open Access
- Author:
- Kakumanu, Akshay
- Graduate Program:
- Integrative Biosciences
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- September 07, 2017
- Committee Members:
- Shaun A Mahony, Dissertation Advisor/Co-Advisor
Shaun A Mahony, Committee Chair/Co-Chair
Benjamin Franklin Pugh, Committee Member
Ross Cameron Hardison, Committee Member
Yu Zhang, Outside Member
Esteban O. Mazzoni, Committee Member - Keywords:
- discriminative features
transcription factor binding sites
DNaseI hypersensitive sites
motif discovery
cellular reprogramming
transcription factor binding
spinal motor neurons
Ngn2
Ascl1
Lhx3
Isl1 - Abstract:
- Transcription factors (TF) bind to highly selective regions of the genome. When ectopically expressed in a defined combination, they enable direct conversion of cellular identities. The thesis of this dissertation is that stratification of TF binding targets through meaningful genomic annotations, followed by the discovery of associated sequence signals, will greatly enhance our understanding of binding site selectivity. However, such analyses are often confounded by uneven overlaps of genomic annotations. As a principle approach to motif discovery, this dissertation introduces SeqUnwinder – a computational framework to deconvolve sequence features associated with overlapping genomic annotations at regulatory sites. Using the novel analyses abilities of SeqUnwinder, we make important contributions towards understanding TF binding in neuronal reprogramming systems. It was previously shown that overexpression of Ngn2, Isl1 and Lhx3 (NIL) effectively programs spinal motor neurons from mouse embryonic stem cells (mESC) within 48 hours. This dissertation unravels the mechanisms of TF binding in NIL reprogramming. Based on SeqUnwinder’s prediction, we show that Onecut factors act as facilitators and synergize with Isl1/Lhx3 to gain access to additional binding targets crucial for reprogramming. In another study, we provide insights into the mechanism through which two pro-neuronal basic helix loop helix factors, Ascl1 and Ngn2, program general neurons with preferential inclinations towards specific neuronal subtypes. Using SeqUnwinder’s approach, we show that the binding preferences of the DNA binding domains of Ascl1 and Ngn2 directly bestows them with distinct instructive abilities to program different neuronal subtypes.