An effort to identify novel proteins in alternative polyadenylation

Thesis Type: Doctorate

Institution Of The Thesis: Middle East Technical University, Graduate School of Natural and Applied Sciences, Turkey

Approval Date: 2020

Thesis Language: English


Supervisor: Ayşe Elif Erson Bensan


Polyadenylation, the addition of a poly (A) tail to a nascent messenger RNA (mRNA), is a tightly regulated process occurring co-transcriptionally. Human pre-mRNA 3’ processing complex is a multi-protein machinery: Core complexes of this machinery can be listed as; i) Cleavage and Polyadenylation Specificity Factor (CPSF), ii) Cleavage Stimulatory Factor (CSTF), and iii) Cleavage Factors Im and IIm (CFIm and CFIIm). Majority of the human genes have multiple poly(A) signals which emphasizes the significance of alternative polyadenylation (APA), which is the regulated selection of alternate polyadenylation signals on mRNAs. APA may change the post-transcriptional fate of a eukaryotic mRNA by affecting its stability, localization and translational activity through altering availability of cis-regulatory elements on mRNA molecule. It is known that proliferative factors alter poly(A) signal selection in normal and cancer cells with a tendency towards increased 3’ UTR shortening. Avoiding negative regulatory trans-factors such as microRNAs, these shortened 3’UTRs have been linked to increased protein levels. These events may explain how certain proteins are upregulated in cancer cells despite the lack of activating mutations. However, it is not yet fully understood how proximal signals are selected in proliferative cells. At this point we hypothesize that, along with core subunits of polyadenylation machinery there may be other proteins which may play a role in selection of poly (A) signals. Aim of this thesis was to identify new proteins that may play a role in APA. For this purpose, we chose proximity-dependent biotin identification coupled with mass spectrometry analysis to identify interacting partners of CSTF2 to better understand how specific poly (A) sites are selected.