Assembling the jigsaw puzzle: CBX2 isoform 2 and its targets in disorders/differences of sex development

Abstract

BackgroundOne of the defining moments of human life occurs early during embryonic development, when individuals sexually differentiate into either male or female. Perturbation of this process can lead to disorders/differences of sex development (DSD). Chromobox protein homolog 2 (CBX2) has two distinct isoforms, CBX2.1 and CBX2.2: the role of CBX2.1 in DSD has been previously established, yet to date the function of the smaller isoform CBX2.2 remains unknown.MethodsThe genomic DNA of two 46,XY DSD patients was analysed using whole exome sequencing. Furthermore, protein/DNA interaction studies were performed using DNA adenine methyltransferase identification (DamID) to identify putative binding partners of CBX2. Finally, in vitro functional studies were used to elucidate the effect of wild‐type and variant CBX2.2 on selected downstream targets.ResultsHere, we describe two patients with features of DSD i.e. atypical external genitalia, perineal hypospadias and no palpable gonads, each patient carrying a distinct CBX2.2 variant, p.Cys132Arg (c.394T>C) and p.Cys154fs (c.460delT). We show that both CBX2.2 variants fail to regulate the expression of genes essential for sexual development, leading to a severe 46,XY DSD defect, likely because of a defective expression of EMX2 in the developing gonad.ConclusionOur study indicates a distinct function of the shorter form of CBX2 and by identifying several of its unique targets, can advance our understanding of DSD pathogenesis and ultimately DSD diagnosis and management.

Disorders of Sex Development—Novel Regulators, Impacts on Fertility, and Options for Fertility Preservation

“…Another CBX2 isoform (CBX2.2) was implicated in the aetiology of 46,XY GD after the identification of deleterious heterozygous variants in two patients with 46,XY partial GD. These CBX2.2 variants failed to regulate the expression of genes essential for gonad development, primarily EMX2 [80]. CBX2 isoforms are a cause of 46,XY GD and to date no information about fertility potential has been reported.…”

CBX2-dependent transcriptional landscape: implications for human sex development and its defects

“…As mentioned, CBX2 has two isoforms, partly distinct in sequence and functions 9,12 . In order to create a Sertoli cell line with a stable CBX2 knockout, durably void of both CBX2 isoforms, a CRISPR/Cas9 construct targeting exon1 was transfected into NT2-D1 cells.…”

“…The clinical importance of CBX2.1 was highlighted by the diagnosis of CBX2 deficiency in 46,XY patients 5,9 . Here, we expand on the previously gained insight by Eid et al .…”

“…CBX2 has a second isoform, CBX2.2, which has very recently been identified as a DSD gene in two unrelated 46,XY patients presenting with complete gonadal dysgenesis 9 . Transactivation studies showed that the variant CBX2.2 failed to regulate the expression of several genes including EMX2, an important factor for bipotential gonadal development.…”

Novas perspectivas no diagnóstico etiológico dos distúrbios do desenvolvimento sexual 46, XY

“…No presente estudo, identificamos uma variante em heterozigose na isoforma 2 (CBX2.2), p.Cys154Alafs*62 em um paciente com disgenesia gonadal parcial. Outra variante nesta mesma isoforma p.Cys132Arg havia sido previamente identificada durante estudo de tese de doutorado da aluna Camila Gomes (131) (132) .…”