African genomes are marked by extensive complexity in the number and distribution of variants, yet remain under-represented in genetic databases and the human reference genome. This gap in representation limits the broad application of genomic medicine. Sickle cell disease (SCD) - one of the most common monogenic diseases - has its highest prevalence in Africa, and variation in disease severity has consistently been linked to the beta-globin locus, including levels of fetal hemoglobin (HbF). Modulation of HbF is central to current SCD gene therapies; however, the inherent complexity and variation at the locus in African genomes presents a challenge to translating these advances to Africa. Here, we align long-read single molecule sequences (LRS) targeted to the beta-globin region to the hg38 and T2T-CHM13v2 genome references in 40 individuals with SCD, predominantly recruited from three African countries. We demonstrate that the expanded T2T-CHM13v2 reference sequence at this locus reduces Structural Variant (SV) calls by 70% and uncovers uncaptured single nucleotide variants (SNVs). Across the cluster we report 343 SVs and 196 SNVs that have not been previously reported, including in LRS data from the All of Us project. By including African populations from ethnolinguistic groups that have not been previously surveyed we improve variant resolution and bolster evidence for observed variation. Finally, we identify a common ∼4kb insertion locus overlapping the HBB promoter among individuals with high HbF. These results demonstrate the utility of combining a comprehensive reference genome with LRS in African populations to uncover genomic variation at disease-associated loci.

BACKGROUND:Hearing impairment (HI) is one of the most common sensory disorders globally. There is limited data on the genetics of HI in Africa. We aim to identify the genetic causes of HI in Cameroon. METHODS:We used whole-exome sequencing (WES) to study the cause of HI in multiplex families from Cameroon. Cell-based experiments using HEK293T assessed the pathogenicity of targeted variants, and a knockout mouse model was used to investigate a candidate gene. RESULTS:Here, we report forty-six multiplex families totaling 106 affected individuals with HI. Most families present with non-syndromic HI (NSHI) (87%, n = 40/46). Causative variants in known genes are identified in 76% (n = 35/46) of families, including nine genes that cause both syndromic HI (SHI) and NSHI, ten that underlie NSHI, and six that cause SHI. MYO15A and TMC1 are the most common genes underlying HI and are observed in six and three families, respectively. Three candidate genes (SUN2, TGM6, and TMC3) are identified. In vitro studies find that a biallelic variant in SUN2 [c.475 G > A:p.(Val159Ile); NM_015374.3] leads to mislocalization and decreased expression in HEK293T cells. In mice, SUN2 protein expression localizes explicitly to the supporting cells of the inner hair cells. Auditory thresholds of the Sun2 knockout mice are comparable to those of wild-type, suggesting that SUN2 is not a credible HI gene. CONCLUSION:This study reports a high diagnostic rate using WES for HI and highlights the opportunity for variant and gene discoveries in African populations. These findings inform the prospects for gene therapy in HI.

Childhood-onset essential hypertension (COEH) is characterized by elevated blood pressure of unknown etiology diagnosed before age 18. COEH affects ~2.1% of children in the United States (U.S.), with a higher prevalence among those of self-reported African ancestries. The genetic underpinnings of COEH are unknown, but its early onset, high heritability, low correlation with environmental factors, and low prevalence suggest a role for monogenic variation. To investigate evidence of Mendelian inheritance in COEH, we performed genome sequencing in 207 individuals from the U.S., including 73 probands and their relatives. We identified six candidate genes with strong evidence of involvement in hypertension in ten families, including SYNE1 (three families), HMCN1 (two), TNS1 (two), FBN2 (one), THBS2 (one), and LRRFIP1 (one). Additionally, six genes with moderate evidence of involvement in hypertension were identified in eight families. Implicated variants in all candidate genes showed evidence of autosomal recessive inheritance, with the resulting candidate genes predominantly expressed in vessels and enriched in individuals of African ancestry. All identified variants were predicted to be damaging to the resulting protein by multiple in silico tools and were either novel or exceedingly rare in population databases. By sequencing an additional 88 probands from South Africa with COEH, we found that 25% (22/88) and 9% (8/88) of them carried at least two putatively damaging variants in SYNE1 and HMCN1 , respectively. Leveraging data from the All of Us initiative, we found that individuals with a history of pediatric essential hypertension are more likely to have damaging variants in our strong candidates as compared to individuals with no history of hypertension in childhood. Notably, SYNE1 , HMCN1 , and TNS1 are presumed to play a role in activating Rho-kinase (ROCK), a protein that regulates contractility in vascular smooth muscle cells (VSMCs). Reduced expression of strong candidates thus far assessed (i.e., SYNE1, HMCN1 ) leads to an abnormal VSMCs phenotype that can be rescued by a downstream ROCK pathway inhibitor. Our findings suggest that genes associated with COEH primarily regulate vascular tone, offering insights that could inform the development of targeted therapeutics and promote precision medicine in the future.

Genetic studies and phenotypic expansion of hearing loss (HL) for people living in Africa are greatly needed. We evaluated the clinical phenotypes of three affected siblings presenting non-syndromic (NS) HL and five unaffected members of a consanguineous Ghanaian family. Analysis of exome sequence data was performed for all affected and one unaffected family members. In-depth genetic and cellular characterization studies were performed to investigate biological significance of the implicated variant using bioinformatic tools and cell-based experimentation. Audiological examinations showed severe-to-profound, bilateral, symmetrical, and post-lingual onset. The whole-exome sequencing (WES) identified a homozygous frameshift variant: MARVEL domain containing 2 (MARVELD2):c.1058dup;p.(Val354Serfs*5) in all affected siblings. This frameshift variant leads to an early stop codon insertion and predicted to be targeted by nonsense medicated decay (mutant protein predicted to lack conserved C-terminal domain if translated). Cell immunofluorescence and immunocytochemistry studies exposed the functional impact of the mutant protein's expression, stability, localization, protein-protein binding, barrier function, and actin cytoskeleton architecture. The identified variant segregates with NSHL in the index Ghanaian family. The data support this nonsense variant as pathogenic, likely to impact the homeostasis of ions, solutes, and other molecules, compromising membrane barrier and signaling in the inner ear spaces.

Childhood-onset essential hypertension (COEH) is an uncommon form of hypertension in which children under the age of 18 have sustained blood pressures greater or equal to the 95th percentile for their age, sex, and height. Childhood blood pressure is known to be a predictor of future blood pressure status; thus, children with COEH are at risk for developing hypertension-related complications as adults, including heart attack, stroke, and renal disease. The heritability of COEH is quoted as between 70-80%, however, the genes and genetic models underlying COEH susceptibility remain unknown and understudied. The relative rarity, high heritability, low correlation with environmental factors, and overrepresentation in African ancestries of COEH suggest a role for rare genetic variation in its pathogenesis. Using genome sequencing, we identified rare and putatively damaging variants in HCMN1 —an essential component of the extracellular matrix architecture in vascular smooth muscle—segregating in an autosomal recessive manner in two families with COEH. To evaluate its functional context, we developed a mouse model of HMCN1 , in which we measured the blood pressure of juvenile Hmcn1 -/- , Hmcn1 +/- , and wild-type mice (4-8 weeks) via non-invasive tail cuffs daily for two weeks, and subsequently measured the same mice as adults via telemetry hourly for two weeks (N=6 per sex, 2 per genotype). Tail cuff data revealed female Hmcn1 -/- mice to have lower blood pressures than female wild-type mice on average (Systolic 126±14 vs. 113±18 mmHg, p>0.05), while male Hmcn1 -/- mice had significantly higher blood pressures than male wild-type mice (Systolic 120±13 vs. 131±12 mmHg, p<0.05). Telemetry data revealed an interactive effect between activity level and genotype on blood pressure—as activity level increased, female Hmcn1 -/- mice had significantly lower blood pressures than wild-type, whereas male Hmcn1 -/- mice had significantly higher blood pressures than wild-type (p<0.01). Post-mortem histology of a female Hmcn1 -/- mouse revealed degeneration and a focal dissecting aneurysm in the aorta. There are currently no published data on the functional impact of HCMN1 on blood pressure or heart function, making our study the first of its kind. Additional studies, specifically those investigating age-related effects on blood pressure in our mouse model, to confirm our findings and understand the pathophysiological effects of HCMN1 loss-of-function on blood pressure are underway.

Childhood -onset essential hypertension (COEH) is an uncommon form of hypertension that manifests in childhood or adolescence and, in the United States, disproportionately affects children of African ancestry. The etiology of COEH is unknown, but its childhood onset, low prevalence, high heritability, and skewed ancestral demography suggest the potential to identify rare genetic variation segregating in a Mendelian manner among affected individuals and thereby implicate genes important to disease pathogenesis. However, no COEH genes have been reported to date. Here, we identify recessive segregation of rare and putatively damaging missense variation in the spectrin domain of spectrin repeat containing nuclear envelope protein 1 ( SYNE1 ), a cardiovascular candidate gene, in 3 of 16 families with early -onset COEH without an antecedent family history. By leveraging exome sequence data from an additional 48 COEH families, 1,700 in-house trios, and publicly available data sets, we demonstrate that compound heterozygous SYNE1 variation in these COEH individuals occurred more often than expected by chance and that this class of biallelic rare variation was significantly enriched among individuals of African genetic ancestry. Using in vitro shRNA knockdown of SYNE1 , we show that reduced SYNE1 expression resulted in a substantial decrease in the elasticity of smooth muscle vascular cells that could be rescued by pharmacological inhibition of the downstream RhoA/Rho-associated protein kinase pathway. These results provide insights into the molecular genetics and underlying pathophysiology of COEH and suggest a role for precision therapeutics in the future.

Background: The Allostatic load score (ALS) is a composite score that measures the cumulative physiological burden of sustained exposure to stress and biological dysregulation. ALS has markers capture risk in 4 domains (neuro-endocrine, metabolic, cardiovascular, and immunological). Higher ALS in adults predict cardiovascular disease (CVD), but it is unknown whether ALS is associated with CVD risk in youth and young adults (YA), and whether the risk and association would differ at such an early age. Aims: In youth and YA we compared 1) the total ALS; 2) the cardiovascular profile according to high and low ALS; and 3) the association of ALS with markers of CVD. Methods: Participants in the African PRospective study on the Early Detection and Identification of Cardiovascular disease and HyperTension (AfricanPREDICT) were 587 youth<24years (48% Male, 21±2 years) and 579 YA≥24years (49% Male, 27±2 years) were evaluated. The ALS was calculated from fasting biomarkers using the upper quartile population distribution of dehydroepiandrosterone (DHEA), cortisol, interleukin-6 (IL-6), C-reactive protein (hsCRP), BMI, systolic-BP, diastolic-BP, and lower quartile of HDL-cholesterol. The upper quartile of ALS in the whole population was defined as high ALS. Retinal vessel analyses, pulse wave velocity, and echocardiography were obtained and associations with ALS determined by regression analyses adjusted for sex, ethnicity, cotinine, gamma-glutamyl-transferase (GGT) and physical activity. Results: ALS median range was 1-8, and a high ALS of >3 was observed in 35% of youth, and 47% YA. In youth with high ALS, ALS associated with narrower retinal arteries (Adj R 2 :0.16, β-coefficient, -1.73, (95%CI), -2.95, -0.94), P =0.052), greater pulse wave velocity (Adj R 2 :0.56, 0.75 (0.48, 1.98), P =0.021), increased left ventricular mass index (Adj R 2 :0.61, 2.21 (0.43, 8.46) P =0.048), lower left ventricular function (global longitudinal strain) (Adj R 2 :0.37, -0.45 (-1.06, -0.06) P =0.015). Similarly, in YA with high ALS, ALS associated with narrower retinal arteries (Adj R 2 :0.43, -4.54 (-6.58, -2.50), P <0.001), greater pulse wave velocity (Adj R 2 :0.44, 0.62 (0.09, 1.17), P =0.043), increased left ventricular mass index (Adj R 2 :0.53, 2.62 (1.52, 5.76) P =0.032), lower diastolic function (E/A ratio) (Adj R 2 :0.27, -0.11 (-0.18, -0.04) P =0.002). Conclusion: Youth and YA with high ALS had a more adverse CVD risk profile and greater association with markers of cardiovascular aging compared to their low ALS counterparts. ALS and allostatic dysregulation should be evaluated in youth and YA, when assessing current and future cardiovascular risk.

The genetic etiology of non-syndromic hearing impairment (NSHI) is highly heterogeneous with over 124 distinct genes identified. The wide spectrum of implicated genes has challenged the implementation of molecular diagnosis with equal clinical validity in all settings. Differential frequencies of allelic variants in the most common NSHI causal gene, gap junction beta 2 (GJB2), has been described as stemming from the segregation of a founder variant and/or spontaneous germline variant hot spots. We aimed to systematically review the global distribution and provenance of founder variants associated with NSHI. The study protocol was registered on PROSPERO, the International Prospective Register of Systematic Reviews, with the registration number "CRD42020198573". Data from 52 reports, involving 27,959 study participants from 24 countries, reporting 56 founder pathogenic or likely pathogenic (P/LP) variants in 14 genes (GJB2, GJB6, GSDME, TMC1, TMIE, TMPRSS3, KCNQ4, PJVK, OTOF, EYA4, MYO15A, PDZD7, CLDN14, and CDH23), were reviewed. Varied number short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs) were used for haplotype analysis to identify the shared ancestral informative markers in a linkage disequilibrium and variants' origins, age estimates, and common ancestry computations in the reviewed reports. Asia recorded the highest number of NSHI founder variants (85.7%; 48/56), with variants in all 14 genes, followed by Europe (16.1%; 9/56). GJB2 had the highest number of ethnic-specific P/LP founder variants. This review reports on the global distribution of NSHI founder variants and relates their evolution to population migration history, bottleneck events, and demographic changes in populations linked with the early evolution of deleterious founder alleles. International migration and regional and cultural intermarriage, coupled to rapid population growth, may have contributed to re-shaping the genetic architecture and structural dynamics of populations segregating these pathogenic founder variants. We have highlighted and showed the paucity of data on hearing impairment (HI) variants in Africa, establishing unexplored opportunities in genetic traits.

This study describes the roles of laboratory information management systems (LIMS) in multi-site genetics studies in Africa. We used the HiGeneS Africa project as a case study. The study participants were recruited in six African countries between 2019 to 2021. The Baobab LIMS, a server–client-based system (an African-led innovation) was used for the coordination of the biospecimen. The development phase of the LIMS showcased the team formation, data collection, biospecimen collection, and shipment strategies. The implementation phase showcased the biospecimen registration, processing, and quality control (QC) analytics. The sample QC was done using Nanodrop, Qubit, and PicoGreen/gDNATapestation assays. The results showed that a total of 3144 study participants were recruited from Cameroon, Ghana, Mali, Rwanda, Senegal, and South Africa. The biospecimen registration provided a comprehensive registry that included patient demographics, genetic information, and clinical and blood/saliva samples from the proband and family relatives. The QC analyzes identified 30 samples that failed QC, linked to overdue storage in the freezer before DNA extraction. The LIMS components implemented in this project formed a structure that can be upscaled to artificial intelligence-based LIMS. In conclusion, this study represents the largest and the most diverse collection of biospecimens for the genetic study of hearing impairment in Africa to date. A well-characterized LIMS should be recommended for multi-site molecular studies, particularly in Africa, to enhance African participation in global genomic medicine.

We investigated hearing impairment (HI) in 51 families from Ghana with at least two affected members that were negative for GJB2 pathogenic variants. DNA samples from 184 family members underwent whole-exome sequencing (WES). Variants were found in 14 known non-syndromic HI (NSHI) genes [26/51 (51.0%) families], five genes that can underlie either syndromic HI or NSHI [13/51 (25.5%)], and one syndromic HI gene [1/51 (2.0%)]. Variants in CDH23 and MYO15A contributed the most to HI [31.4% (16/51 families)]. For DSPP, an autosomal recessive mode of inheritance was detected. Post-lingual expression was observed for a family segregating a MARVELD2 variant. To our knowledge, seven novel candidate HI genes were identified (13.7%), with six associated with NSHI (INPP4B, CCDC141, MYO19, DNAH11, POTEI, and SOX9); and one (PAX8) with Waardenburg syndrome. MYO19 and DNAH11 were replicated in unrelated Ghanaian probands. Six of the novel genes were expressed in mouse inner ear. It is known that Pax8-/- mice do not respond to sound, and depletion of Sox9 resulted in defective vestibular structures and abnormal utricle development. Most variants (48/60; 80.0%) have not previously been associated with HI. Identifying seven candidate genes in this study emphasizes the potential of novel HI genes discovery in Africa.

Hearing impairment (HI) is a silent planetary health crisis that requires attention worldwide. The prevalence of HI in South Africa is estimated as 5.5 in 100 live births, which is about 5 times higher than the prevalence in high-income countries. This also offers opportunity to drive progressive science, technology and innovation policy, and health systems. We present here a systematic analysis and review on the prevalence, etiologies, clinical patterns, and genetics/genomics of HI in South Africa. We searched PubMed, Scopus, African Journals Online, AFROLIB, and African Index Medicus to identify the pertinent studies on HI in South Africa, published from inception to April 30, 2021, and the data were summarized narratively. We screened 944 records, of which 27 studies were included in the review. The age at diagnosis is ∼3 years of age and the most common factor associated with acquired HI was middle ear infections. There were numerous reports on medication toxicity, with kanamycin-induced ototoxicity requiring specific attention when considering the high burden of tuberculosis in South Africa. The Waardenburg Syndrome is the most common reported syndromic HI. The Usher Syndrome is the only syndrome with genetic investigations, whereby a founder mutation was identified among black South Africans (MYO7A-c.6377delC). GJB2 and GJB6 genes are not major contributors to nonsyndromic HI among Black South Africans. Furthermore, emerging data using targeted panel sequencing have shown a low resolution rate in Black South Africans in known HI genes. Importantly, mutations in known nonsyndromic HI genes are infrequent in South Africa. Therefore, whole-exome sequencing appears as the most effective way forward to identify variants associated with HI in South Africa. Taken together, this article contributes to the emerging field of planetary health genomics with a focus on HI and offers new insights and lessons learned for future roadmaps on genomics/multiomics and clinical studies of HI around the world.

We have previously reported CLIC5A and SLC12A2 variants in two families from Cameroon and Ghana, segregating non-syndromic hearing impairment (NSHI). In this study, biological assays were performed to further functionally investigate the pathogenicity of CLIC5 [c.224T>C; p.(L75P)] and SCL12A2 [c.2935G>A: p.(E979K)] variants. Ectopic expression of the proteins in a cell model shows that compared to wild-type, both the CLIC5A and SLC12A2 variants were overexpressed. The mutant CLIC5A protein appears as aggregated perinuclear bodies while the wild-type protein was evenly distributed in the cytoplasm. Furthermore, cells transfected with the wild-type CLIC5A formed thin membrane filopodia-like protrusions which were absent in the CLIC5A mutant expressing and control cells. On the other hand, the wild-type SLC12A2 expressing cells had an axon-like morphology which was not observed in the mutant expressing and control cells. A network analysis revealed that CLIC5A can interact with at least eight proteins at the base of the stereocilia. This study has generated novel biological data associated with the pathogenicity of targeted variants in CLIC5A and SLC12A2, found in two African families, and therefore expands our understanding of their pathobiology in hearing impairment.

Congenital hearing impairment (HI) is genetically heterogeneous making its genetic diagnosis challenging. Investigation of novel HI genes and variants will enhance our understanding of the molecular mechanisms and to aid genetic diagnosis. We performed exome sequencing and analysis using DNA samples from affected members of two large families from Ghana and Pakistan, segregating autosomal-dominant (AD) non-syndromic HI (NSHI). Using in silico approaches, we modeled and evaluated the effect of the likely pathogenic variants on protein structure and function. We identified two likely pathogenic variants in SLC12A2 , c.2935G>A:p.(E979K) and c.2939A>T:p.(E980V), which segregate with NSHI in a Ghanaian and Pakistani family, respectively. SLC12A2 encodes an ion transporter crucial in the homeostasis of the inner ear endolymph and has recently been reported to be implicated in syndromic and non-syndromic HI. Both variants were mapped to alternatively spliced exon 21 of the SLC12A2 gene. Exon 21 encodes for 17 residues in the cytoplasmatic tail of SLC12A2, is highly conserved between species, and preferentially expressed in cochlear tissues. A review of previous studies and our current data showed that out of ten families with either AD non-syndromic or syndromic HI, eight (80%) had variants within the 17 amino acid residue region of exon 21 (48 bp), suggesting that this alternate domain is critical to the transporter activity in the inner ear. The genotypic spectrum of SLC12A2 was expanded and the involvement of SLC12A2 in ADNSHI was confirmed. These results also demonstrate the role that SLC12A2 plays in ADNSHI in diverse populations including sub-Saharan Africans.

Approximately half of congenital hearing impairment cases are inherited, with non-syndromic hearing impairment (NSHI) being the most frequent clinical entity of genetic hearing impairment cases. A family from Cameroon with NSHI was investigated by performing exome sequencing using DNA samples obtained from three family members, followed by direct Sanger sequencing in additional family members and controls participants. We identified an autosomal dominantly inherited novel missense variant [NM_001174116.2:c.918G>T; p.(Q306H)] in DMXL2 gene (MIM:612186) that co-segregates with mild to profound non-syndromic sensorineural hearing impairment . The p.(Q306H) variant which substitutes a highly conserved glutamine residue is predicted deleterious by various bioinformatics tools and is absent from several genome databases. This variant was also neither found in 121 apparently healthy controls without a family history of hearing impairment , nor 112 sporadic NSHI cases from Cameroon. There is one previous report of a large Han Chinese NSHI family that segregates a missense variant in DMXL2. The present study provides additional evidence that DMXL2 is involved in hearing impairment etiology, and we suggest DMXL2 should be considered in diagnostic hearing impairment panels.

Physiologically, the human and murine hearing systems are very similar, justifying the extensive use of mice in experimental models for hearing impairment (HI). About 340 murine HI genes have been reported; however, whether variants in all human-mouse ortholog genes contribute to HI has been rarely investigated. In humans, nearly 120 HI genes have been identified to date, with GJB2 and GJB6 variants accounting for half of congenital HI cases, of genetic origin, in populations of European and Asian ancestries, but not in most African populations. The contribution of variants in other known genes of HI among the populations of African ancestry is poorly studied and displays the lowest pick-up rate. We used whole exome sequencing (WES) to investigate pathogenic and likely pathogenic (PLP) variants in 34 novel human-mouse orthologs HI genes, in 40 individuals from Cameroon and South Africa diagnosed with non-syndromic hearing impairment (NSHI), and compared the data to WES data of 129 ethnically matched controls. In addition, protein modeling for selected PLP gene variants, gene enrichment, and network analyses were performed. A total of 4/38 murine genes, d6wsu163e, zfp719, grp152 and minar2, had no human orthologs. WES identified three rare PLP variants in 3/34 human-mouse orthologs genes in three unrelated Cameroonian patients, namely: OCM2, c.227G>C p.(Arg76Thr) and LRGI1, c.1657G>A p.(Gly533Arg) in a heterozygous state, and a PLP variant MCPH1, c.2311C>G p.(Pro771Ala) in a homozygous state. In silico functional analyses suggest that these human-mouse ortholog genes functionally co-expressed interactions with well-established HI genes: GJB2 and GJB6. The study found one homozygous variant in MCPH1, likely to explain HI in one patient, and suggests that human-mouse ortholog variants could contribute to the understanding of the physiology of hearing in humans.

Background: This study aimed to gain an understanding of the challenges faced by people with hearing impairment (HI) in Cameroon, their understanding of the causes of HI, and how challenges could be remedied to improve the quality of life of persons with HI. Methods: Semi-structured one-on-one in-depth interviews and observation of participant behaviour when answering questions were used to collect data from 10 HI professionals (healthcare workers and educationists), and 10 persons affected by HI (including caregivers). Results: The results show that the different groups associate the causes of HI to genetics, environmental factors, and a spiritual curse. There were reported cases of stigma and discrimination of persons with HI, with people sometimes referring to HI as an “intellectual disorder.” Interviewees also highlighted the difficulty persons with HI have in accessing education and healthcare services and suggested the need for the government and health researchers to develop strategies for the prevention and early diagnosis of HI. These strategies include (1) the awareness of the general population regarding HI, (2) the development of facilities for the proper management and new-born screening of HI, and (3) the implementation of a premarital screening to reduce the burden of HI of genetic origin. Conclusions: This study confirms the difficult social interaction and access to proper management faced by persons with HI in Cameroon and further highlights the need to educate populations on the causes of HI for a better acceptance of individuals with HI in the Cameroonian society.

Hearing impairment (HI) is a sensory disorder with a prevalence of 0.0055 live births in South Africa. DNA samples from a South African family presenting with progressive, autosomal dominant non-syndromic HI were subjected to whole-exome sequencing, and a novel monoallelic variant in REST [c.1244GC; p.(C415S)], was identified as the putative causative variant. The co-segregation of the variant was confirmed with Sanger Sequencing. The variant is absent from databases, 103 healthy South African controls, and 52 South African probands with isolated HI. In silico analysis indicates that the p.C415S variant in REST substitutes a conserved cysteine and results in changes to the surrounding secondary structure and the disulphide bonds, culminating in alteration of the tertiary structure of REST. Localization studies using ectopically expressed GFP-tagged Wild type (WT) and mutant REST in HEK-293 cells show that WT REST localizes exclusively to the nucleus; however, the mutant protein localizes throughout the cell. Additionally, mutant REST has an impaired ability to repress its known target AF1q. The data demonstrates that the identified mutation compromises the function of REST and support its implication in HI. This study is the second report, worldwide, to implicate REST in HI and suggests that it should be included in diagnostic HI panels.