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Proper repair of damaged DNA is critical for the maintenance of genome stability. A complex composed of Integrator subunit 3 (Ints3), single-stranded DNA-binding protein 1 (SSB1), and SSB-interacting protein 1 (SSBIP1) is required for efficient homologous recombination-dependent repair of double-strand breaks (DSBs) and ataxia-telangiectasia mutated (ATM)-dependent signaling pathways. It is known that in this complex the Ints3 N-terminal domain scaffolds SSB1 and SSBIP1. However, the molecular basis for the function of the Ints3 C-terminal domain remains unclear. Here, we present the crystal structure of the Ints3 C-terminal domain, uncovering a HEAT-repeat superhelical fold. Using structure and mutation analysis, we show that the C-terminal domain exists as a stable dimer. A basic groove and a cluster of conserved residues on two opposite sides of the dimer bind single-stranded RNA/DNA (ssRNA/ssDNA) and Integrator complex subunit 6 (Ints6), respectively. Dimerization is required for nucleic acid binding, but not for Ints6 binding. Additionally, in vitro experiments using HEK 293T cells demonstrate that Ints6 interaction is critical for maintaining SSB1 protein level. Taken together, our findings establish the structural basis of a multifunctional Ints3 C-terminal module, allowing us to propose a novel mode of nucleic acid recognition by helical repeat protein and paving the way for future mechanistic studies.

SOSS1 is a single-stranded DNA (ssDNA)-binding protein complex that plays a critical role in double-strand DNA break (DSB) repair. SOSS1 consists of three subunits: INTS3, SOSSC, and hSSB1 with INTS3 serving as a scaffold to stabilize this complex. Moreover, the integrator complex subunit 6 (INTS6) participates in the DNA damage response through direct binding to INTS3 but how INTS3 interacts with INTS6 thereby, impacting DBS repair is not clear. Here, we determined the crystal structure of the C-terminus of INTS3 (INTS3c) in complex with the C-terminus of INTS6 (INTS6c) at a resolution of 2.4 Å. Structure analysis revealed that two INTS3c subunits dimerize and interact with INTS6c via conserved residues. Subsequent biochemical analyses confirmed that INTS3c forms a stable dimer and INTS3 dimerization is important for recognizing the longer ssDNA. Perturbation of INTS3c dimerization and disruption of the INTS3c/INTS6c interaction, impair the DSB repair process. Altogether, these results unravel the underappreciated role of INTS3 dimerization and the molecular basis of INTS3/INTS6 interaction in DSB repair.

In this study, we aimed to determine whether the pseudogene integrator complex subunit 6 pseudogene 1 (INTS6P1) in plasma could be used as a novel approach to screen for and detect hepatocellular carcinoma (HCC). We explored the clinical role of INTS6P1: First, the expression level of INTS6P1 was measured in a cohort of 33 HCC tissue samples and adjacent normal liver tissue, next, the INTS6P1 expression was detected in the culture medium and tumor cells in a cellular experiment, and last, the diagnostic performance of INTS6P1 was examined in an independent cohort of 100 people. The expression level of INTS6P1 was remarkably downregulated in the HCC tissues compared with that in the normal liver tissues (p = 0.0066). In plasma, the INTS6P1 levels were significantly decreased in HCC patients compared with non-HCC patients (p

Human single-stranded DNA-binding protein 1 (hSSB1) plays an important role in the DNA damage response and the maintenance of genomic stability. It has been shown that the core hSSB1 complex contains hSSB1, INTS3 and C9orf80. Using protein affinity purification, we have identified integrator complex subunit 6 (INTS6) as a major subunit of the core hSSB1 complex. INTS6 forms a stable complex with INTS3 and hSSB1 both in vitro and in vivo. In this complex, INTS6 directly interacts with INTS3. In response to the DNA damage response, along with INTS3 and hSSB1, INTS6 relocates to the DNA damage sites. Moreover, the hSSB1–INTS complex regulates the accumulation of RAD51 and BRCA1 at DNA damage sites and the correlated homologous recombination.

Review on INTS6 (integrator complex subunit 6), with data on DNA, on the protein encoded, and where the gene is implicated.