טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentTzunz-Henig Noa
SubjectNovel Role for the Transcription Factor Sfp1 in mRNA
Decay
DepartmentDepartment of Medicine
Supervisor Professor Mordechai Choder
Full Thesis textFull thesis text - English Version


Abstract

Gene expression consists of several processes, each represent a distinct stage. Recently, it has become clear that consecutive stages are mechanistically coupled. Gene expression, therefore, can be viewed as a continuous process. However, little is known about coupling between processes occurring in different cellular compartments.

Rpb4p, a subunit of RNA polymerase II complex that synthesizes all mRNAs, is the only protein known to be involved both in mRNA synthesis and in mRNA decay. It is specifically required for efficient decay of a class of mRNAs encoding Protein Biosynthetic Factors (PBF).

PBF genes are transcribed by several transcription factors including Sfp1 and Fhl1. Several observations provoked us to examine whether Sfp1 is involved in Rpb4-mediated mRNA decay:

1. Sfp1 and Rpb4 form two hybrid interactions.

2. Both shuttle between the nucleus and the cytoplasm.

3. Cellular localization of both proteins is similarly responsive to stress conditions.

4. Both proteins are associated with poly-ribosomes and probably regulate translation.

We first determined decay kinetics, and found that efficient decay of PBF mRNAs is dependent upon SFP1. In contrast, decay of ACT1 mRNA, a non-PBF mRNA, was independent on SFP1. Moreover, GFP-Sfp1p accumulated in cytoplasmic foci that behaved like genuine P-bodies, where mRNA degradation is executed, suggesting that Sfp1 is directly involved in mRNA decay in the cytoplasm.

To examine whether Sfp1 functions in mRNA decay in the context of other proteins, we examined another transcription factor of ribosomal genes - Fhl1p. Lack of FHL1 also affected specifically the decay kinetics of PBF mRNAs, leading to enhanced decay of these mRNAs.

The involvement of Sfp1 and Rpb4 in synthesis and decay of PBF mRNAs led us to hypothesize that these proteins act together in post-transcriptional roles.

Consistently, we found that efficient export of GFP-Rpb4 was dependent upon SFP1.

Moreover, we observed a high rate of co-localization between Sfp1-bodies and Rpb4- bodies, implying that these proteins act together while mRNAs are in P-bodies.

To conclude, Sfp1 regulates both the synthesis and, as we show here, also the decay of some PBF mRNAs. This is the first report that a specific transcription factor is involved also in the decay of the respective mRNAs. However, the exact role of Sfp1 and Fhl1 in mRNA decay remains to be determined.

More generally, Sfp1 may be a component of a larger complex that links the synthesis and decay of class-specific mRNAs, thus controlling their appropriate level in the cytoplasm.