ADrosophilaXPD model links cell cycle coordination with neuro-development and suggests links to cancer

Abstract

XPD functions in transcription, DNA repair and in cell cycle control. Mutations in human XPD (also known as ERCC2) mainly cause three clinical phenotypes: xeroderma pigmentosum (XP), Cockayne syndrome (XP/CS) and trichothiodystrophy (TTD), and only XP patients have a high predisposition to developing cancer. Hence, we developed a fly model to obtain novel insights into the defects caused by individual hypomorphic alleles identified in human XP-D patients. This model revealed that the mutations that displayed the greatest in vivo UV sensitivity in Drosophila did not correlate with those that led to tumor formation in humans. Immunoprecipitations followed by targeted quantitative MS/MS analysis showed how different xpd mutations affected the formation or stability of different transcription factor IIH (TFIIH) subcomplexes. The XP mutants most clearly linked to high cancer risk, Xpd R683W and R601L, showed a reduced interaction with the core TFIIH and also an abnormal interaction with the Cdk-activating kinase (CAK) complex. Interestingly, these two XP alleles additionally displayed high levels of chromatin loss and free centrosomes during the rapid nuclear division phase of the Drosophila embryo. Finally, the xpd mutations showing defects in the coordination of cell cycle timing during the Drosophila embryonic divisions correlated with those human mutations that cause the neurodevelopmental abnormalities and developmental growth defects observed in XP/CS and TTD patients.

Mms19 is a mitotic gene that permits Cdk7 to be fully active as a Cdk-activating kinase

“…Our model therefore proposes that Mms19 binding releases Xpd from core TFIIH by releasing it from p44 and that it additionally releases the trimeric CAK complex from Xpd, allowing it to become an active Cdk-activating kinase. Interestingly, the regions on Xpd needed for the binding of the three different proteins have been mapped and they seem to explain such a competition model (summarized by Stettler et al, 2014 ). The region in XPD spanning amino acids 277-286 is essential for its interaction with MMS19 ( Vashisht et al, 2015 ) and it is part of the ARCH domain (amino acids 245-443), which is also required for its stable association with the trimeric CAK complex ( Sandrock and Egly, 2001 ).…”

“…The region in XPD spanning amino acids 277-286 is essential for its interaction with MMS19 ( Vashisht et al, 2015 ) and it is part of the ARCH domain (amino acids 245-443), which is also required for its stable association with the trimeric CAK complex ( Sandrock and Egly, 2001 ). Similarly, the binding sites for Mms19 and p44 are at least adjacent if not overlapping ( Sandrock and Egly, 2001 ; Dubaele et al, 2003 ; Stettler et al, 2014 ). Steric interference and direct competition for binding sites might therefore provide the structural basis for the exclusive interaction of XPD with either Mms19 or the CAK (and TFIIH) complex.…”

A CK1 FRET biosensor reveals that DDX3X is an essential activator of CK1ε

“…Immunoprecipitates were resolved by SDS-PAGE and gel bands sent for mass spectrometry. Mass spectrometry data acquisition and analysis (Program SpC PepShaker) was performed at the Mass Spectrometry and Proteomics Laboratory (Department of Clinical Research, University of Bern) essentially as described in Stettler et al (2014) .…”

Nucleotide excision repair genes shaping embryonic development

“…Transgenic flies carrying human-like alleles with mutations reported in human patients reproduce these defects, suggesting that Drosophila is a good model for these studies [107]. Another existing model for another TFIIH subunit, XPD, has been reported in Drosophila , allowing for different human mutations to be tested during development [108]. This Drosophila model revealed an Xpd function in cell cycle coordination which is affected by XP/CS and TTD mutations [108].…”