Structural analysis of the CIC-HMG-ETV5-DNA complex

Abstract

Capicua (CIC) acts as a transcriptional repressor in fly and mammals and it is conserved from C.elegans to human. Previous research shows that its function is also conserved as regulator of downstream genes of the RTK (Receptor Tyrosine Kinase) signaling pathway. CIC represses tll, hkb and zen which is related to dorsoventral patterning and development of terminal region in fly and ETV1, 4 and 5 in mammal. It is known that CIC recognizes the octameric target site, TGAATGAA at the promoter region of target genes through the DNA binding region, the HMG-box domain. CIC is related to generation of several diseases including one of the polyQ diseases, spinocerebellar ataxia type 1 (SCA1), and several cancers. Somatic mutations found in the HMG-box DNA binding domain in CIC have been implicated with several cancers such as oligodendroglioma, oligoastrocytoma, and adenocarcinoma. Especially, the majority of patients with oligodendroglioma possesses mutations in CIC and the number of mutations were identified in the HMG-box domain. However, the molecular basis of the DNA binding of CIC and the effect of the somatic mutations found in cancers on DNA binding have not been investigated. Here, we report the crystal structure of the HMG-box domain of CIC complexed with its target DNA, the promoter of Ets Translocation Variant 5 (ETV5). The crystal structure was solved at $3.0\angstrom\$ in space group P32. The structure shows that the HMG-box domain posits a roughly L-shaped structure and recognizes the minor groove leading to DNA bending. We examined the interactions between HMG-box domain and ETV5-DNA observed in crystal structure in solution by electrophoretic mobility shift assay (EMSA). Also, our structure combined with result of EMSA revealed that mutations in the HMG-box domain found in the cancers abrogate the interaction with DNA. These results provide the molecular insight into the DNA binding of CIC and the effect of carcinogenic mutations on the DNA binding.