G·A·B·B·A

ASH1 mRNA translation has been shown to be required for efficient anchoring of ASH1 mRNA to
the bud-tip during the process of mRNA localization.
Khd1p, a 3-KH domain containing protein, has been shown to be involved in ASH1 mRNA
localization and has been proposed to be a translation repressor (Irie et al., 2002). This thesis
focuses on the role of Khd1p in ASH1 mRNP architecture, specially in its relationship to other
members of the locasome. Results presented show that Khd1p is a part of an RNA-independent
protein complex that includes She3p and She2p, indicating Khd1p is a part of the locasome via
protein-protein interactions. Furthermore, the observation that Khd1p and She3p interact
independently of She2p indicates that another protein complex exists and that this complex could be
another cargo for the SHE mRNA localization machinery. On the other hand, results presented in
this work show that Khd1p floats with ER membranes, as observed for She2p (Schmid et al., 2006)
indicating that Khd1p is also implicated in ER-associated. The fact that Khd1p floats independently
of She2p indicates that it might be involved in ER inheritance, via a direct binding to She3p. During
the course of this work, it was shown that Khd1p is phosphorylated by Yck1p at the cellular
periphery and upon phosphorylation, Khd1p releases the ASH1 mRNA translation block (Paquin et
al., 2007) thus proving the model for Khd1p function in translation repression during ASH1 mRNA
transport and localization.
Khd1p, an hnRNP K homolog, has also been shown to play a role in telomeric silencing and be
bound to subtelomeric chromatin (Denisenko et al., 2002). This thesis addresses how Khd1p, an
mRNA binding protein, can also play a role in this process. Results presented show that Khd1p
contributes to telomericgene transcriptional silencing. Associated to loss of KHD1 I found that
Sir2p levels are downregulated, leading to loss of silencing in mutants known to be silenced (e.g.
RPD3 – Rundlett et al., 1996). This loss of silencing could be partially rescued with wildtype copies
of SIR2.
Results presented in this work suggest that KHD1 is also involved in binding to the telomerase
RNA subunit, TLC1 or even in its biogenesis.
In addition, a loss of KHD1 also led to increased sensitivity to DNA damaging agents (UV and
MMS) of rad52 and xrs2 null mutants. Results presented show that KHD1 contributes to NHEJ
DNA repair efficiency and that this loss of NHEJ efficiency can be rescued with a wildtype copy of
KHD1. In addition, a point mutant I183R, cannot rescue this NHEJ defect, thus implying that this
residue or the folding of this KH domain is essential for NHEJ function.

Results presented in this thesis establish Khd1p as a putative new cargo for the SHE machinery and
also as an ER associated protein. Furthermore, results presented here also establish Khd1p as a
regulator of gene transcriptional telomeric silencing, associated to a downregulation of Sir2p. In
addition, we show that Khd1p contributes to NHEJ DNA repair, indicating a much wider action
than just in mRNA localization.