INTS13Mutations Causing a Developmental Ciliopathy Disrupt Integrator Complex Assembly

Abstract

Oral-facial-digital syndromes (OFD) are a heterogeneous group of congenital disorders characterized by malformations of the face and oral cavity, and digit anomalies. To date, mutations in 12 ciliary-related genes have been identified that cause several types of OFD, suggesting that OFDs constitute a subgroup of developmental ciliopathies. Through homozygosity mapping and exome sequencing of two families with variable OFD type 2, we identified distinct germline mutations in INTS13, a subunit of the Integrator complex. This 14-component complex associates with RNAPII and can cleave nascent RNA to modulate gene expression. We determined that INTS13 utilizes a discrete domain within its C-terminus to bind the Integrator cleavage module, which is disrupted by the identified germline INTS13 mutations. Depletion of INTS13 disrupts ciliogenesis in human cultured cells and causes dysregulation of a broad collection of ciliary genes. Accordingly, its knockdown in Xenopus embryos lead to motile cilia anomalies. Altogether, we show that mutations in INTS13 cause an autosomal recessive ciliopathy, which reveals key interactions within Integrator components.

Structure of the catalytic core of the Integrator complex

“…Our findings suggests that the Integrator complex is highly modular, not unlike the related CPSF/CPF complex ( Casañal et al., 2017 ; Zhang et al., 2020b ). Furthermore, we provide evidence that two of the modules (INTS4/9/11 and INTS10/13/14) interact with each other ( Figure S1 ), consistent with recent reports ( Mascibroda et al., 2020 ; Sabath et al., 2020 ). Notably, other INTSs were also detected in our experiment, but their abundance was much lower compared with the three complexes discussed.…”

Mechanistic basis of gene-specific transcription regulation by the Integrator complex

“…55 Consistent with our data, previously published interactomes of ZMYND8, ZEB1, LEC subunit ICE1, and mouse ZNF609, contain several INTS in addition to INTS10-13-14, 36,51, [56][57][58] and conversely numerous INTS interactomes also feature ZMYND8, ZNF609, and ZNF655. 35,37,47,[59][60][61][62][63] Together these data indicate that the regulatory factors interact with the whole INT by directly binding the INTS10-13-14 module and that this function is conserved to vertebrates (Supplemental Figure 2B). We therefore propose that INTS10-13-14 form the INT TF binding module (ITFM).…”

“…19,32,[35][36][37][38][39] Consistent with a crucial function during cell differentiation and stimulus response, INTS deletions are embryonically lethal and INTS mutations cause developmental disorders, many of which manifest with neurological disease components. 37,[40][41][42][43] Despite the clear observation that transcription attenuation via INT is required for diverse biological response pathways, it is unclear how the pivotal decision between productive elongation and abortive early termination is made in the correct spatiotemporal and genespecific manner. Here, we show that several sequence-specific transcription factors (TFs), and transcription effector complexes (chromatin remodeler INO80; little elongation complex) bind INT directly on two conserved surfaces of the INTS10-13-14 module.…”

Sparse dictionary learning recovers pleiotropy from human cell fitness screens

“…Finally, Webster inferred the fitness effect of perturbing the INTS10–13-14 heterotrimer, a physical and functional module of the Integrator complex ( Barbieri et al, 2018 ; Mascibroda et al, 2020 ; Pfleiderer and Galej, 2021 ; Sabath et al, 2020 ). INTS10, INTS14, and INTS13 were the strongest loadings on this function (1.1, 0.9, 0.8, respectively), along with a fourth gene, C7orf26 (1.0).…”