Gutenberg Open Science
The Open Science Repository of Johannes Gutenberg University Mainz.
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Recent Submissions
Die Hemisphärenlateralisation im Altersvergleich – eine fMRT-Studie
(2024) Bulheller, Theresa Elisabeth; Tüscher, Oliver
Deciphering the role of USH1G/SANS in proteinprotein interactions and nuclear shuttling.
(2025) Fritze, Jacques Sebastian; Wolfrum, Uwe
In my thesis I investigated the molecular mechanisms underlying Human Usher
syndrome (USH), particularly focusing on the protein USH1G/SANS. Usher syndrome,
a clinically and genetically heterogeneous disorder that leads to hearing and vision
loss. The USH1G gene encodes the scaffold protein SANS, which is highly expressed
in the eye and ear. Recently, SANS has also been found in the cell nucleus, where it
participates in the regulation of pre-mRNA splicing.
In my thesis I aimed to elucidate the interactions of SANS with splicing-related
proteins PRPF31 and PRPF6, as well as the mechanism of SANS’ nuclear-cytoplasmic
shuttling. Additionally, I established a method to monitor the nuclear transfer of
PRPF31 by live cell imaging.
I show that SANS interacts with PRPF31 and PRPF6 at specific sites within its
CENTn domain. Using FRET-based interaction assays and AlphaFold2, we identified
for PRPF31 and PRPF6 specific binding sites in the CENTn domain of SANS, namely
binding to the structured CENTn1 and to the unstructured CENTn2, respectively. In
addition, we found evidence for sequential binding of PRPF31 and PRPF6 to the SANS
molecule, which might be crucial for role of SANS in splicing processes in the nucleus.
To fulfill its nuclear functions, SANS needs to be transported into the nucleus. In
my thesis I identified two nuclear localization sequences (NLSs) and two nuclear export
sequences (NESs). I highlighted their critical roles in SANS subcellular localization and
in nuclear–cytoplasmic shuttling. Comparative analysis with SANS’ paralogue
ANKS4B revealed distinct localization patterns, with ANKS4B more confined to the
cytoplasm due to a lack of NLS.
Using pathogenic variants of USH1G/SANS, we demonstrated altered interactions
with the splicing proteins PRPF31 and PRPF6. Additionally, these pathogenic variants
displayed subcellular mislocalization, likely impacting SANS’ function in splicing
regulation and in the development of USH.
Further, I provide data on the nuclear mobility of photoactivatable fluorescencetagged
PRPF31 and the rapid transfer of splicing components between nuclear
compartments. With these results, I could lay the groundwork for future studies on
SANS-dependent molecular dynamics.
Collectively, I enlighten the role of SANS in the nucleus. My findings advance the
understanding of the molecular functions of SANS and its broader implications for
USH1G pathogenesis.