Penetrance Of Phenotype Research Articles

AN ATYPICAL PRESENTATION OF FAMILIAL MEDITERRANEAN FEVER

<h3>Introduction</h3> Familial Mediterranean Fever(FMF) is thought to be autosomal recessive disease characterized by periodic fevers associated with serositis. In this case we present a patient with FMF with only one mutation in MEFV gene, suggesting a more variable penetrance of the FMF phenotype. <h3>Case Description</h3> An 8-year-old female diagnosed with juvenile idiopathic arthritis and asymptomatic hematuria presented for elevated immunoglobulins and inflammatory markers. She had no history of recurrent/severe infections or fevers. She endorsed monoarticular joint pains and abdominal pains. Family history was significant for a 1-year-old sister diagnosed with PFAPA and a 15-months-old brother who died from human metapneumovirus associated pulmonary hemorrhage. On exam, she appeared well-nourished, no rash/oral ulcers/abdominal tenderness. Labs were significant for CRP-54, ESR-52, IgG-1950, IgA-327, IgM-230. Urinalysis had 3+ blood without proteinuria. Taken together, her history raised concern for monogenic inborn error of immunity and a primary immunodeficiency panel was sent. Her panel revealed a heterozygous pathogenic variance of MEFV(V726A) responsible for FMF. Her father was later also diagnosed with FMF responsive to colchicine with the same pathogenic MEFV variant. Her cytokines were elevated, including IL-1B, and she was subsequently started on colchicine 0.6mg BID and canakinumab 75mg q4-weeks. Following treatment, she had normalization of inflammatory markers (Table 1) and resolution of joint pains. <h3>Discussion</h3> This case highlights the existence of a subset of FMF patients who have only one mutation in the MEFV gene. Broadening our understanding of the complex genetics and variable penetrance of FMF can prevent delays in diagnosis and treatment.

The transcriptional co-repressor SEED DORMANCY 4-LIKE (AtSDR4L) promotes the embryonic-to-vegetative transition in Arabidopsis thaliana.

Repression of embryonic traits during the seed-to-seedling phase transition requires the inactivation of master transcription factors associated with embryogenesis. How the timing of such inactivation is controlled is unclear. Here, we report on a novel transcriptional co-repressor, Arabidopsis thaliana SDR4L, that forms a feedback inhibition loop with the master transcription factors LEC1 and ABI3 to repress embryonic traits post-imbibition. LEC1 and ABI3 regulate their own expression by inducing AtSDR4L during mid to late embryogenesis. AtSDR4L binds to sites upstreamof LEC1 and ABI4, and these transcripts are upregulated in Atsdr4l seedlings. Atsdr4l seedlings phenocopy a LEC1 overexpressor. The embryonic traits of Atsdr4l can be partially rescued by impairing LEC1 or ABI3. The penetrance and expressivity of the Atsdr4l phenotypes depend on both developmental and external cues, demonstrating the importance of AtSDR4L in seedling establishment under suboptimal conditions.

Reclassification of a likely pathogenic Dutch founder variant in KCNH2; implications of reduced penetrance.

Variants in KCNH2, encoding the human ether a-go-go (hERG) channel that is responsible for the rapid component of the cardiac delayed rectifier K+ current (IKr), are causal to long QT syndrome type 2 (LQTS2). We identified eight index patients with a new variant of unknown significance (VUS), KCNH2:c.2717C > T:p.(Ser906Leu). We aimed to elucidate the biophysiological effect of this variant, to enable reclassification and consequent clinical decision-making. A genotype-phenotype overview of the patients and relatives was created. The biophysiological effects were assessed independently by manual-, and automated calibrated patch clamp. HEK293a cells expressing (i) wild-type (WT) KCNH2, (ii) KCNH2-p.S906L alone (homozygous, Hm) or (iii) KCNH2-p.S906L in combination with WT (1:1) (heterozygous, Hz) were used for manual patching. Automated patch clamp measured the variants function against known benign and pathogenic variants, using Flp-In T-rex HEK293 KCNH2-variant cell lines. Incomplete penetrance of LQTS2 in KCNH2:p.(Ser906Leu) carriers was observed. In addition, some patients were heterozygous for other VUSs in CACNA1C, PKP2, RYR2 or AKAP9. The phenotype of carriers of KCNH2:p.(Ser906Leu) ranged from asymptomatic to life-threatening arrhythmic events. Manual patch clamp showed a reduced current density by 69.8 and 60.4% in KCNH2-p.S906L-Hm and KCNH2-p.S906L-Hz, respectively. The time constant of activation was significantly increased with 80.1% in KCNH2-p.S906L-Hm compared with KCNH2-WT. Assessment of KCNH2-p.S906L-Hz by calibrated automatic patch clamp assay showed a reduction in current density by 35.6%. The reduced current density in the KCNH2-p.S906L-Hz indicates a moderate loss-of-function. Combined with the reduced penetrance and variable phenotype, we conclude that KCNH2:p.(Ser906Leu) is a low penetrant likely pathogenic variant for LQTS2.

Arginase 1 deficiency presenting as complicated hereditary spastic paraplegia.

Argininemia or arginase deficiency is a metabolic disorder caused by pathogenic variants in ARG1 and consists of a variable association of progressive spastic paraplegia, intellectual disability, and seizures. Hereditary spastic paraplegia (HSP) is a group of inherited diseases whose main feature is a progressive gait disorder characterized by lower limb spasticity. This study presents 7 patients with arginase 1 deficiency from 6 different families, all with an initial diagnosis of complicated HSP. We evaluated the clinical data of 7 patients belonging to six independent families who were diagnosed with hyperargininemia in a neurogenetics outpatient clinic. All patients had lower limb spasticity and six had global developmental delay. Five individuals had intellectual disability and two had epilepsy. Psychiatric abnormalities were seen in two patients. In two participants of this study, MRI disclosed thinning of the corpus callosum. Molecular diagnosis was made by whole exome sequencing. All variants were present in homozygosis; we identified two novel missense variants, one novel frameshift variant, and one previously published missense variant. Clinical diagnosis of early onset complicated hereditary spastic paraplegia was made in all patients. Two patients were initially suspected of having SPG11 due to thinning of the corpus callosum. As argininemia may present with a highly penetrant phenotype of spastic paraplegia associated with additional symptoms, this disease may represent a specific entity amongst the complicated HSPs.

Genome Sequencing and Transcriptome Profiling in Twins Discordant for Mayer-Rokitansky-Küster-Hauser Syndrome.

To identify potential genetic causes for Mayer-Rokitansky-Küster-Hauser syndrome (MRKH), we analyzed blood and rudimentary uterine tissue of 5 MRKH discordant monozygotic twin pairs. Assuming that a variant solely identified in the affected twin or affected tissue could cause the phenotype, we identified a mosaic variant in ACTR3B with high allele frequency in the affected tissue, low allele frequency in the blood of the affected twin, and almost absent in blood of the unaffected twin. Focusing on MRKH candidate genes, we detected a pathogenic variant in GREB1L in one twin pair and their unaffected mother showing a reduced phenotypic penetrance. Furthermore, two variants of unknown clinical significance in PAX8 and WNT9B were identified. In addition, we conducted transcriptome analysis of affected tissue and observed perturbations largely similar to those in sporadic cases. These shared transcriptional changes were enriched for terms associated with estrogen and its receptors pointing at a role of estrogen in MRKH pathology. Our genome sequencing approach of blood and uterine tissue of discordant twins is the most extensive study performed on twins discordant for MRKH so far. As no clear pathogenic differences were detected, research to evaluate other regulatory layers are required to better understand the complex etiology of MRKH.

Molecular and behavioral consequences of Ube3a gene overdosage in mice.

Chromosome 15q11.2-q13.1 duplication syndrome (Dup15q syndrome) is a severe neurodevelopmental disorder characterized by intellectual disability, impaired motor coordination, and autism spectrum disorder. Chromosomal multiplication of the UBE3A gene is presumed to be the primary driver of Dup15q pathophysiology, given that UBE3A exhibits maternal monoallelic expression in neurons and that maternal duplications typically yield far more severe neurodevelopmental outcomes than paternal duplications. However, studies into the pathogenic effects of UBE3A overexpression in mice have yielded conflicting results. Here, we investigated the neurodevelopmental impact of Ube3a gene overdosage using bacterial artificial chromosome-based transgenic mouse models (Ube3aOE) that recapitulate the increases in Ube3a copy number most often observed in Dup15q. In contrast to previously published Ube3a overexpression models, Ube3aOE mice were indistinguishable from wild-type controls on a number of molecular and behavioral measures, despite suffering increased mortality when challenged with seizures, a phenotype reminiscent of sudden unexpected death in epilepsy. Collectively, our data support a model wherein pathogenic synergy between UBE3A and other overexpressed 15q11.2-q13.1 genes is required for full penetrance of Dup15q syndrome phenotypes.

Variable paralog expression underlies phenotype variation.

Human faces are variable; we look different from one another. Craniofacial disorders further increase facial variation. To understand craniofacial variation and how it can be buffered, we analyzed the zebrafish mef2ca mutant. When this transcription factor encoding gene is mutated, zebrafish develop dramatically variable craniofacial phenotypes. Years of selective breeding for low and high penetrance of mutant phenotypes produced strains that are either resilient or sensitive to the mef2ca mutation. Here, we compared gene expression between these strains, which revealed that selective breeding enriched for high and low mef2ca paralog expression in the low- and high-penetrance strains, respectively. We found that mef2ca paralog expression is variable in unselected wild-type zebrafish, motivating the hypothesis that heritable variation in paralog expression underlies mutant phenotype severity and variation. In support, mutagenizing the mef2ca paralogs, mef2aa, mef2b, mef2cb, and mef2d demonstrated modular buffering by paralogs. Specifically, some paralogs buffer severity while others buffer variability. We present a novel, mechanistic model for phenotypic variation where variable, vestigial paralog expression buffers development. These studies are a major step forward in understanding the mechanisms of facial variation, including how some genetically resilient individuals can overcome a deleterious mutation.

Circulating miRNA-16 in inflammatory bowel disease and some clinical correlations - a cohort study in Bulgarian patients.

The pathogenesis of the inflammatory bowel disease (IBD) includes chronic inflammation and altered immune reactions. There are several publications, reporting that micro ribonucleic acids (miRNAs) may serve as a diagnostic biomarker with a potential to assess inflammation severity and treatment response1,2 in IBD patients. The objective of the study is to assess and correlate the serum expression of circulating miR-16 in IBD patients with some clinical parameters, such as extent, activity and severity of the disease. 70 IBD patients [35 with ulcerative colitis (UC) and 35 with Crohn's disease (CD)] were included in the study. Serum miR-16 expression in both IBD diseases was assessed using reverse transcription quantitative real time PCR (RT-qPCR). Circulating miR-16 levels were also correlated with disease extent, activity and severity indices [Crohn's Disease Activity Index (CDAI), Montreal classification, Partial Mayo score]. Serum expression of miR-16 in the 70 patients was also compared to miR-16 serum levels in 30 healthy control subjects. The patients' group showed mean serum miR-16 expression of 3.07 for CD, 1.97 for UC and 1.61 for the control group of healthy subjects with a significant difference in the expression between groups. There is a significant correlation between increased serum expression of miR-16 and disease activity, extent and severity. The increased miR-16 serum expression correlates with disease activity, intestinal localization of CD, stenotic and penetrating phenotype. MiR-16 could serve as a potential biomarker to assess inflammation.

Development and Validation of the PREMMplus Model for Multigene Hereditary Cancer Risk Assessment.

With the availability of multigene panel testing (MGPT) for hereditary cancer risk assessment, clinicians need to assess the likelihood of pathogenic germline variants (PGVs) across numerous genes in parallel. This study's aim was to develop and validate a clinical prediction model (PREMMplus) for MGPT risk assessment. PREMMplus was developed in a single-institution cohort of 7,280 individuals who had undergone MGPT. Logistic regression models with Least Absolute Shrinkage and Selection Operator regularization were used to examine candidate predictors (age, sex, ethnicity, and personal/family history of 18 cancers/neoplasms) to estimate one's likelihood of carrying PGVs in 19 genes (broadly categorized by phenotypic overlap and/or relative penetrance: 11 category A [APC, BRCA1/2, CDH1, EPCAM, MLH1, MSH2, MSH6, biallelic MUTYH, PMS2, and TP53] and eight category B genes [ATM, BRIP1, CDKN2A, CHEK2, PALB2, PTEN, RAD51C, and RAD51D]). Model performance was validated in nonoverlapping data sets of 8,691 and 14,849 individuals with prior MGPT ascertained from clinic- and laboratory-based settings, respectively. PREMMplus (score ≥ 2.5%) had 93.9%, 91.7%, and 89.3% sensitivity and 98.3%, 97.5%, and 97.8% negative-predictive value (NPV) for identifying category A gene PGV carriers in the development and validation cohorts, respectively. PREMMplus assessment (score ≥ 2.5%) had 89.9%, 85.6%, and 84.2% sensitivity and 95.0%, 93.5%, and 93.5% NPV, respectively, for identifying category A/B gene PGV carriers. Decision curve analyses support MGPT for individuals predicted to have ≥ 2.5% probability of a PGV. PREMMplus accurately identifies individuals with PGVs in a diverse spectrum of cancer susceptibility genes with high sensitivity/NPV. Individuals with PREMMplus scores ≥ 2.5% should be considered for MGPT.

MicroRNA-101a enhances trabecular bone accrual in male mice

High-throughput microRNA sequencing was performed during differentiation of MC3T3-E1 osteoblasts to develop working hypotheses for specific microRNAs that control osteogenesis. The expression data show that miR-101a, which targets the mRNAs for the epigenetic enzyme Ezh2 and many other proteins, is highly upregulated during osteoblast differentiation and robustly expressed in mouse calvaria. Transient elevation of miR-101a suppresses Ezh2 levels, reduces tri-methylation of lysine 27 in histone 3 (H3K27me3; a heterochromatic mark catalyzed by Ezh2), and accelerates mineralization of MC3T3-E1 osteoblasts. We also examined skeletal phenotypes of an inducible miR-101a transgene under direct control of doxycycline administration. Experimental controls and mir-101a over-expressing mice were exposed to doxycycline in utero and postnatally (up to 8 weeks of age) to maximize penetrance of skeletal phenotypes. Male mice that over-express miR-101a have increased total body weight and longer femora. MicroCT analysis indicate that these mice have increased trabecular bone volume fraction, trabecular number and trabecular thickness with reduced trabecular spacing as compared to controls. Histomorphometric analysis demonstrates a significant reduction in osteoid volume to bone volume and osteoid surface to bone surface. Remarkably, while female mice also exhibit a significant increase in bone length, no significant changes were noted by microCT (trabecular bone parameters) and histomorphometry (osteoid parameters). Hence, miR-101a upregulation during osteoblast maturation and the concomitant reduction in Ezh2 mediated H3K27me3 levels may contribute to the enhanced trabecular bone parameters in male mice. However, the sex-specific effect of miR-101a indicates that more intricate epigenetic mechanisms mediate physiological control of bone formation and homeostasis.

Variation in Susceptibility to Permethrin in Culex pipiens and Culex restuans Populations in the Great Lakes Region of the United States.

Two Culex pipiens form Pipiens colony strains and a field population of Cx. restuans from Michigan were susceptible and a Cx. pipiens form Molestus colony strain was comparatively less susceptible to a dose of 43 μg/ml of permethrin in Centers for Disease Control and Prevention (CDC) bottle bioassays. Using this diagnostic dose and these populations as controls, adult female Cx. pipiens and Cx. restuans were reared from egg rafts from 28 sites in Illinois, Ohio, Michigan, Iowa, Indiana, Wisconsin, and Minnesota. Tested mosquitoes showed high mortality in populations from 12 sites, less mortality (90-96%) at 9 sites, and less than 90% mortality from 7 sites during 30-min exposures. However, all tested populations showed 97-100% mortality at 60 min, indicating low phenotypic penetrance of resistance factors. These results indicate variation in susceptibility to permethrin in populations of West Nile virus vectors in the Great Lakes region of the United States, with evidence of modest resistance at 7 of 28 (25%) of the sampled populations.

P793: SCREENING OF NATURALLY OCCURRING CXCR4 VARIANTS FOR IDENTIFICATION OF NOVEL PATHOGENIC MUTATIONS FOR WHIM SYNDROME

Background: WHIM (Warts, Hypogammaglobulinemia, Infections, Myelokathexis) syndrome is a rare, autosomal-dominant primary immunodeficiency marked by neutropenia and lymphopenia. Classically, WHIM syndrome pathogenesis is causally linked to a variety of heterozygous gain-of-function mutations in the C-terminus of chemokine receptor CXCR4, a master regulator of immune cell trafficking and homeostasis. As of February 2022, 18 variants in CXCR4 have been implicated in WHIM syndrome, while a substantial number remain unexplored for their association with the disease. Using in vitro functional assays, we previously found that impaired receptor internalization and enhanced chemotaxis are conserved in all CXCR4WHIM variants and that defective internalization correlates with neutropenia, the most penetrant phenotype of patients with WHIM syndrome. We used this approach to confirm functional defects in primary cells isolated from patients harboring a novel variant, suggesting that the in vitro profiling approach can potentially drive identification of pathogenic variants. Aims: We aimed to functionally characterize multiple previously uncharacterized variants of CXCR4 using in vitro assays. We also examined allele frequencies of these novel variants to estimate the potential number of individuals harboring these variants with a long-term goal of determining the actual prevalence of WHIM syndrome. Methods: The CXCR4-negative K562 cell line was used as a model system to express 53 novel CXCR4 variants identified in patient and population databases (ClinVar, Ensembl, CentoMD, GnomAD) and genetic screening initiatives (Invitae PATH4WARD) found in the C-terminus (hotspot of CXCR4WHIM mutations) as well as throughout the protein. The effects of these mutations on CXCR4 internalization and chemotaxis were studied in cells stimulated with the C-X-C chemokine ligand 12 (CXCL12). The in vitro functional parameters were compared with known pathogenic CXCR4 variants and were used to assign a cumulative score (impaired internalization + enhanced chemotaxis) of functional defect severity. Results: Out of 53 selected variants, 42 were missense (ms), 6 were frameshift (fs), 3 were nonsense (ns), and 2 were transcript variants (tv). Eighteen of the screened variants led to a decreased internalization of CXCR4 in comparison to the wild-type (WT) receptor upon stimulation with CXCL12 while 20 variants demonstrated enhanced chemotaxis of cells toward CXCL12. Overall, 34 variants showed defects in at least one CXCR4-dependent marker. Correlation analysis and scoring of the functional parameters in cells expressing mutated or CXCR4WT revealed 17 variants (9 ms, 5 fs, 2 ns, 1 tv) that co-segregated with the known pathogenic WHIM variants. Finally, white blood cell counts in 3 patients harboring novel variants showed that CXCR4 internalization defects in vitro correlated with the severity of neutropenia. Summary/Conclusion: This is the first study characterizing functional impairments in naturally occurring CXCR4 variants via a pipeline of assays intended to predict pathogenicity. Seventeen CXCR4 variants were identified (several outside the C-terminus of CXCR4) that caused in vitro functional defects resembling those exhibited by known WHIM variants. Many (10/17) of these newly identified variants are present at high frequencies (4.4 × 10-4–8.0 × 10-6) in genomic databases. Potential association of these variants with WHIM syndrome is under investigation and is likely to expand current estimates of WHIM syndrome prevalence.

Prophylactic Antitubercular Therapy Is Associated With Accelerated Disease Progression in Patients With Crohn's Disease Receiving Anti-TNF Therapy: A Retrospective Multicenter Study.

INTRODUCTION:Prophylactic antitubercular therapy (ATT) is widely prescribed in patients with Crohn's disease (CD) receiving antitumor necrosis factor (anti-TNF) treatment. However, antitubercular agents have been demonstrated to possess profibrotic effects. We aimed to evaluate whether ATT accelerated disease progression in patients with CD receiving anti-TNF treatment.METHODS:A retrospective, multicenter study was performed in CD patients presented with inflammatory behavior (B1) and treated with anti-TNF agents. Disease progression was defined as the development of a stricturing (B2) or penetrating (B3) phenotype. ATT users were propensity score-matched with non-ATT users. Survival and multivariable Cox analyses were used to identify factors associated with disease progression.RESULTS:We enrolled 441 patients, including 295 ATT users and 146 non-ATT users, with a median follow-up of 3.15 years (interquartile range: 1.6–4.7). The cumulative rates of disease progression in the ATT group were constantly higher than those in the non-ATT group after 1-, 3-, 5-, and 10-year follow-ups, respectively (P = 0.031). Multivariable Cox analysis identified ATT as an independent risk factor for disease progression using both the whole (hazard ratio = 2.22; 95% confidence interval: 1.11–4.48; P = 0.025) and propensity score-matched cohorts (hazard ratio = 2.35; 95% confidence interval: 1.07–5.14; P = 0.033). In subgroup analysis, patients receiving ATT ≥4.5 months had a significantly higher rate of disease progression compared with patients receiving ATT <4.5 months (P = 0.005) and non-ATT treatment (P = 0.036).DISCUSSION:Prophylactic ATT with duration over 4.5 months was associated with disease progression in patients with CD receiving anti-TNF treatment.

Frequency of truncating FLCN variants and Birt-Hogg-Dubé–associated phenotypes in a health care system population

Penetrance estimates of Birt-Hogg-Dubé syndrome (BHD)-associated cutaneous, pulmonary, and kidney manifestations are based on clinically ascertained families. In a health care system population, we used a genetics-first approach to estimate the prevalence of pathogenic/likely pathogenic (P/LP) truncating variants in FLCN, which cause BHD, and the penetrance of BHD-related phenotypes. Exomes from 135,990 patient-participants in Geisinger's MyCode cohort were assessed for P/LP truncating FLCN variants. BHD-related phenotypes were evaluated from electronic health records. Association between P/LP FLCN variants and BHD-related phenotypes was assessed using Firth's logistic regression. P/LP truncating FLCN variants were identified in 35 individuals (1 in 3234 unrelated individuals), 68.6% of whom had BHD-related phenotype(s), including cystic lung disease (65.7%), pneumothoraces (17.1%), cutaneous manifestations (8.6%), and kidney cancer (2.9%). A total of 4 (11.4%) individuals had prior clinical BHD diagnoses. In this health care population, the frequency of P/LP truncating FLCN variants is 60 times higher than the previously reported prevalence. Although most variant-positive individuals had BHD-related phenotypes, a minority were previously clinically diagnosed, likely because cutaneous manifestations, pneumothoraces, and kidney cancer were observed at lower frequencies than in clinical cohorts. Improved clinical recognition of cystic lung disease and education concerning its association with FLCN variants could prompt evaluation for BHD.

A spectrum of recessiveness among Mendelian disease variants in UK Biobank

Recent work has found increasing evidence of mitigated, incompletely penetrant phenotypes in heterozygous carriers of recessive Mendelian disease variants. We leveraged whole-exome imputation within the full UK Biobank cohort (n ∼ 500K) to extend such analyses to 3,475 rare variants curated from ClinVar and OMIM. Testing these variants for association with 58 quantitative traits yielded 102 significant associations involving variants previously implicated in 34 different diseases. Notable examples included a POR missense variant implicated in Antley-Bixler syndrome that associated with a 1.76 (SE 0.27) cm increase in height and an ABCA3 missense variant implicated in interstitial lung disease that associated with reduced FEV1/FVC ratio. Association analyses with 1,134 disease traits yielded five additional variant-disease associations. We also observed contrasting levels of recessiveness between two more-common, classical Mendelian diseases. Carriers of cystic fibrosis variants exhibited increased risk of several mitigated disease phenotypes, whereas carriers of spinal muscular atrophy alleles showed no evidence of altered phenotypes. Incomplete penetrance of cystic fibrosis carrier phenotypes did not appear to be mediated by common allelic variation on the functional haplotype. Our results show that many disease-associated recessive variants can produce mitigated phenotypes in heterozygous carriers and motivate further work exploring penetrance mechanisms.

Dual nicotinic acetylcholine receptor subunit gene knockouts reveal limits to functional redundancy

The nicotinic acetylcholine receptor (nAChR) subunit gene family consists of ten members in Drosophila melanogaster. The mature nAChR is a pentamer assembled from these subunits. Despite recent advances in the in vitro expression of some receptor subunit combinations (nAChR subtypes), the in vivo combinations and stoichiometry of these subtypes remains poorly defined. In addition, there are many potential nAChR signalling roles for different subtypes in insect behaviour, development and physiology. Prior work has shown that nAChR subunit mutants can display altered sleep and mating behaviour, disrupted hormone signalling and reduced locomotion, climbing ability and longevity. Teasing out the specific receptor subunits that are involved in these different functions is potentially made more difficult given that the structural similarity between members of gene families often means that there is a degree of functional redundancy. In order to circumvent this, we created a dual knockout strain for the Dα1 and Dβ2 nAChR subunit genes and examined four traits including insecticide resistance. These subunits had been previously implicated in the response to a neonicotinoid insecticide, imidacloprid. The use of the dual knockout revealed that Dα1 and Dβ2 subunits are involved in signalling that leads to the inflation of wings following adult emergence from the pupal case. The Dβ1 subunit had previously been implicated as a contributor to this function. The lack of a phenotype or low penetrance of the phenotype in the Dα1 and Dβ2 single mutants compared to the dual knockout suggests that these subunits are, to some extent, functionally redundant. We also observed stronger reductions in climbing ability and longevity in the dual knockout. Our findings demonstrate that a dual knockout approach to examining members of the nAChR subunit gene family may increase the power of genetic approaches linking individual subunits and combinations thereof to particular biological functions. This approach will be valuable as the nAChRs are so widely expressed in the insect brain that they are likely to have many functions that hereto remain undetected.

Mutation, selection, and the prevalence of the Caenorhabditis elegans heat-sensitive mortal germline phenotype.

Caenorhabditis elegans strains with the heat-sensitive mortal germline phenotype become progressively sterile over the course of a few tens of generations when maintained at temperatures near the upper range of C. elegans’ tolerance. Mortal germline is transgenerationally heritable, and proximately under epigenetic control. Previous studies have suggested that mortal germline presents a relatively large mutational target and that mortal germline is not uncommon in natural populations of C. elegans. The mortal germline phenotype is not monolithic. Some strains exhibit a strong mortal germline phenotype, in which individuals invariably become sterile over a few generations, whereas other strains show a weaker (less penetrant) phenotype in which the onset of sterility is slower and more stochastic. We present results in which we (1) quantify the rate of mutation to the mortal germline phenotype and (2) quantify the frequency of mortal germline in a collection of 95 wild isolates. Over the course of ∼16,000 meioses, we detected one mutation to a strong mortal germline phenotype, resulting in a point estimate of the mutation rate UMrt≈ 610−5/genome/generation. We detected no mutations to a weak mortal germline phenotype. Six out of 95 wild isolates have a strong mortal germline phenotype, and although quantification of the weak mortal germline phenotype is inexact, the weak mortal germline phenotype is not rare in nature. We estimate a strength of selection against mutations conferring the strong mortal germline phenotype ≈0.1%, similar to selection against mutations affecting competitive fitness. The appreciable frequency of weak mortal germline variants in nature combined with the low mutation rate suggests that mortal germline may be maintained by balancing selection.

Causative Variants for Inherited Cardiac Conditions in a Southeast Asian Population Cohort.

Variable penetrance and late-onset phenotypes are key challenges for classifying causal as well as incidental findings in inherited cardiac conditions. Allele frequencies of variants in ancestry-specific populations, along with clinical variant analysis and interpretation, are critical to determine their true significance. Here, we carefully reviewed and classified variants in genes associated with inherited cardiac conditions based on a population whole-genome sequencing cohort of 4810 Singaporeans representing Southeast Asian ancestries. Eighty-nine (1.85%) individuals carried either pathogenic or likely pathogenic variants across 25 genes. Forty-six (51.7%) had variants in causal genes for familial hyperlipidemia, but there were also recurrent variants in SCN5A and MYBPC3, causal genes for inherited arrhythmia and cardiomyopathy, which, despite previous reports, we determined to lack criteria for pathogenicity. Our findings highlight the incidence of disease-related variants in inherited cardiac conditions and emphasize the value of large-scale sequencing in specific ancestries. Follow-up detailed phenotyping and analysis of pedigrees are crucial because assigning pathogenicity will significantly affect clinical management for individuals and their family members.

P699 Machine Learning-based Prediction for Early Progression in Korean Crohn’s disease: Results from the IMPACT Study

Abstract Background Transcriptome-wide association studies (TWAS) improve to detect functionally relevant loci by leveraging expression quantitative trait loci (eQTLs) from reference panels in relevant tissues. Herein, we developed machine learning-based prediction models using a novel Korean TWAS model for early progression to stricturing or penetrating phenotypes in Korean patients with Crohn’s disease (CD). Methods A total of 431 patients diagnosed with CD were retrospectively enrolled from 15 referral hospitals in Korea. Single-nucleotide polymorphism genotype was analyzed using the Korea Biobank Array. Using the GTEx genotype and Korean chip genotype database from the terminal ileum, a novel Korean TWAS model was developed and compared with a European TWAS model. Predictive models for early progression were trained and cross-validated using logistic regression models and leave-one-out cross validation. Early progression was defined as transition to stricturing or penetrating phenotypes within 2 years from the diagnosis of CD. Results Among the study population, early progression to stricturing and penetrating phenotype was detected in 60 (13.9%) and 73 patients (16.9%), respectively. Combined clinical-gene expression prediction models predicted early progression to stricturing (AUROC, 0.816) and penetrating phenotypes (AUROC, 0.801) more accurately than the models using clinical parameters alone. The following gene expression was significantly associated with early progression: CCDC154, FAM189A2, TAS2R19, FCSK, SP1, and KCNIP1 for stricturing phenotype and PUS7, CCDC146, MLXIP, LRGUK, UROS, and TAFA1 for penetrating phenotype. Conclusion The clinical-genetic machine learning models predicted progression to stricturing or penetrating phenotypes in the early stage of CD. Use of the comprehensive prediction models might enable clinicians to select patients requiring early aggressive therapeutic strategy to prevent disease-related complications.

P119 Hereditary Colorectal Cancer Syndromes and Inflammatory Bowel Diseases: an ECCO CONFER Multicenter Case Series

Abstract Background Hereditary colorectal cancer syndromes (HCCS) are rare polyposis or nonpolyposis syndromes with a higher risk of developing colorectal cancer (CRC). Coexisting inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn’s disease (CD), with HCCS is exceedingly rare and presumably increases the risk of early-onset colorectal cancer. Methods This was a multicenter case series performed as a part of the European Crohn’s and Colitis Organisation (ECCO) Collaborative Network of Exceptionally Rare case reports (CONFER) project. Results This report includes 26 patients with IBD (10 UC, 15 CD, and one with IBD unclassified (IBD-U)) and concomitant HCCS. Among these 26 patients (median age 33 years, IQR 20–44), 15 (57.7%) were males, 24 (92.3%) Caucasians and 2 (7.7%) of Arab origin. Among patients with UC, 4 (40%) had proctitis, 3 (30%) left-sided colitis and 3 (30%) an extensive disease; while, among patients with CD, 8 (53.3%) had ileal, 5 (33.3%) ileocolonic, 2 (13.3%) colonic disease. One of these patients had ileocolonic disease associated with upper disease location and six (40%) had an associated perianal disease. Many of patients with CD had a non-stricturing non-penetrating phenotype (53.3%, n=8), while 20.0% (n=3) and 26.7% (n=4) had a stricturing or penetrating phenotype, respectively. HCCS was diagnosed before the IBD diagnosis in 11 patients (42.3%), after diagnosis of IBD in 11 patients (42.3%) and concurrently in 4 patients (15.4%). Sixteen patients had Lynch syndrome, 7 had FAP, 2 had AFAP and one had MAP. The most frequent genetic mutations were those of APC (n=7) and MLH1 (n= 7). Thirteen patients were treated with immunomodulatory (IMM) therapy. During IMM therapy, only 5 patients developed drug-induced side-effects which did not require drug interruption. One patient treated with vedolizumab developed CRC, despite yearly colonoscopy surveillance, after 4 years of treatment. Overall, CRC developed in 38.5% of patients (n=10): in 5 patients (50%) after IBD diagnosis, in 4 (40%) patients before IBD diagnosis, and in 1 patient the 2 conditions were diagnosed simultaneously. Eighteen (69.2%) patients underwent colectomy or abdominal surgery: 9 patients due to CRC diagnosis, 7 patients preventively due to the underlying HCCS and 2 patients because of active IBD disease. On follow-up, one patient died due to CRC. Conclusion This case series describes the largest series to date of patients with IBD patients and HCCS, with special emphasis on the use of IMM in this population. The most frequent diagnosis of HCCS associated with IBD was Lynch syndrome. These data demonstrate the high malignancy rate and surgical intervention rate in this IBD cohort. The optimal medical approach still needs to be addressed.