טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentErez Neta
SubjectIdentification of Ub/UbL-Related Genes that Maintain the
Differentiated Identity of Adult Drosophila Gut
DepartmentDepartment of Medicine
Supervisor Professor Amir )Oryan )Orian
Full Thesis textFull thesis text - English Version


Abstract

Differentiated cells require continuous and active mechanisms to maintain the differentiated identity and to prevent cells from activating non-relevant gene programs. Identity supervision is crucial to tissue homeostasis and serves as a barrier to tumorigenesis, spontaneous de-differentiation and forced reprogramming. Yet, the mechanisms and genetic networks that maintain the differentiated state are relatively unknown.

An excellent adult tissue for identity studies is the Drosophila intestine and specifically the midgut. It is a highly dynamic tissue that is most similar to the vertebrate and human intestine. Therefore, it represents a powerful model system to study the genetic basis and mechanisms involved in adult tissue homeostasis and is highly relevant to cancer biology. The adult Drosophila gut offers the use of powerful genetic tools, cell-specific marking, conditional manipulation of genes and lineage tracing. It is also optimal for imaging, cutting edge genomic studies and RNAi screening in vivo.

My broad aim in this study was to identify genes and mechanisms that are required for maintaining the differentiated identity of the gut enterocytes using Drosophila melanogaster adult midgut as a model system. I focused on regulators that belong to the ubiquitin and ubiquitin-like pathways. First, I conducted a large-scale RNAi screen and identified seventeen candidate genes whose loss in enterocytes results in loss of cell identity. Next, I focused on Nonstop, a de-ubiquitinase enzyme, and characterized its function as an enterocyte-identity regulator. Lineage tracing of enterocytes established that Nonstop-deficient enterocytes lose their differentiated identity and fail to express enterocyte-specific genes including the differentiated type-A Lamin, Lamin C. In addition, enterocytes that lack Nonstop ectopically express stem cell and non-relevant gene programs. I found that Nonstop silences non-relevant genes indirectly via the expression of Lamin C, that represses stem cell and non-relevant gene-programs. Nonstop is also required for large-scale nuclear organization. RNA-sequencing and ATAC-sequencing profiling established that to maintain identity Nonstop controls enterocyte-specific gene programs by maintaining a unique nucleosome positioning and thereby chromatin configuration. Moreover, the function of Nonstop as an enterocyte-identity regulator is independent of the SAGA complex, but likely involves E(y)2. In sum, my results reveal a potential novel Nonstop-identity protein complex, consisting of Nonstop, E(y)2 and likely other proteins including GAGA factor/Trl, that maintains enterocyte-specific gene signature and safeguards enterocyte identity.