Gene regulation by alternative cleavage and polyadenylation (ApA)
My lab studies gene regulation during development and cancer, and we focus on the regulation and functional consequences of ApA using genome-wide approaches.
DNA is transcribed into the primary transcript. This transcript needs to be processed into a mature messenger RNA (mRNA), because only mature mRNAs can be translated into protein. mRNA processing consists of capping, splicing, and cleavage at the 3' end with subsequent addition of a poly(A) tail. The recognition of ApA sites leads to the generation of mRNA isoforms that differ in the length of their 3' untranslated regions (3'UTRs). The alternative mRNAs produce the wild-type protein but differ in the presence of regulatory elements located in the 3'UTR, such as localization signals, AU-rich elements, or microRNA binding sites. We showed for specific genes that the shorter mRNA isoforms produce on average tenfold more protein. This suggests that ApA is a gene regulatory mechanism to control protein expression levels.
Half of all human genes generate mRNA isoforms with more than one 3'UTR isoform (multi-UTR genes). We developed a next-generation sequencing method, called 3'-seq, to quantify 3'UTR isoform expression. In close collaboration with Christina Leslie’s lab, we established a computational method that allows us to identify differentially expressed 3'UTR isoforms between samples. We investigated samples from diverse human tissues and found that alternative 3'UTR isoform expression is highly tissue specific. About two-thirds of all multi-UTR genes show a significantly different 3'UTR isoform expression pattern in a least two samples. We call these genes polyadeno-regulated multi-UTR (pAM) genes. During differentiation or transformation only a fraction of the multi-UTR genes change their relative isoform expression. But the genes that change their isoform ratios are highly specific to each perturbation. These genes are involved in tissue-specific processes despite being transcribed ubiquitously.
With our current projects we are seeking to answer the following questions:
- What are the differences between ApA in normal cells and cancer cells? What is the role of ApA during the development of cancer? Can a change in ApA pattern mimic genetic aberrations such as amplifications
- What are the differences between ApA in ES cells and differentiated cells? Is a change in ApA isoform expression necessary for differentiation?
- Is a shift toward increased expression of shorter 3'UTR isoforms mostly due to increased recognition of the proximal pA site or is it due to decreased stability of the longer 3'UTR isoforms?
- How do protein levels correlate genome-wide with differences in ApA isoform expression?
- What are the differences between short-term and long-term changes in ApA?
- How is ApA regulated? What is the influence of the promoter, transcription elongation, and 3'UTR elements on the strength of a pA site?
- How does ApA contribute to heterogeneity within a cell population?