SEMA6B variants cause intellectual disability and alter dendritic spine density and axon guidance.

Intellectual disability (ID) is a neurodevelopmental disorder frequently caused by monogenic defects. In this study, we collected 14 SEMA6B heterozygous variants in 16 unrelated patients referred for ID to different centers. Whereas, until now, SEMA6B variants have mainly been reported in patients with progressive myoclonic epilepsy, our study indicates that the clinical spectrum is wider and also includes non-syndromic ID without epilepsy or myoclonus. To assess the pathogenicity of these variants, selected mutated forms of Sema6b were overexpressed in Human Embryonic Kidney 293T (HEK293T) cells and in primary neuronal cultures. shRNAs targeting Sema6b were also used in neuronal cultures to measure the impact of the decreased Sema6b expression on morphogenesis and synaptogenesis. The overexpression of some variants leads to a subcellular mislocalization of SEMA6B protein in HEK293T cells and to a reduced spine density owing to loss of mature spines in neuronal cultures. Sema6b knockdown also impairs spine density and spine maturation. In addition, we conducted in vivo rescue experiments in chicken embryos with the selected mutated forms of Sema6b expressed in commissural neurons after knockdown of endogenous SEMA6B. We observed that expression of these variants in commissural neurons fails to rescue the normal axon pathway. In conclusion, identification of SEMA6B variants in patients presenting with an overlapping phenotype with ID and functional studies highlight the important role of SEMA6B in neuronal development, notably in spine formation and maturation and in axon guidance. This study adds SEMA6B to the list of ID-related genes.

The duplication 17p13.3 phenotype: analysis of 21 families delineates developmental, behavioral and brain abnormalities, and rare variant phenotypes.

Chromosome 17p13.3 is a gene rich region that when deleted is associated with the well-known Miller-Dieker syndrome. A recently described duplication syndrome involving this region has been associated with intellectual impairment, autism and occasional brain MRI abnormalities. We report 34 additional patients from 21 families to further delineate the clinical, neurological, behavioral, and brain imaging findings. We found a highly diverse phenotype with inter- and intrafamilial variability, especially in cognitive development. The most specific phenotype occurred in individuals with large duplications that include both the YWHAE and LIS1 genes. These patients had a relatively distinct facial phenotype and frequent structural brain abnormalities involving the corpus callosum, cerebellar vermis, and cranial base. Autism spectrum disorders were seen in a third of duplication probands, most commonly in those with duplications of YWHAE and flanking genes such as CRK. The typical neurobehavioral phenotype was usually seen in those with the larger duplications. We did not confirm the association of early overgrowth with involvement of YWHAE and CRK, or growth failure with duplications of LIS1. Older patients were often overweight. Three variant phenotypes included cleft lip/palate (CLP), split hand/foot with long bone deficiency (SHFLD), and a connective tissue phenotype resembling Marfan syndrome. The duplications in patients with clefts appear to disrupt ABR, while the SHFLD phenotype was associated with duplication of BHLHA9 as noted in two recent reports. The connective tissue phenotype did not have a convincing critical region. Our experience with this large cohort expands knowledge of this diverse duplication syndrome.

A Novel SPAST/SPG4 Splice-Site Variant in a Family with Dominant Hereditary Spastic Paraplegia.

Some causes of spastic paraplegia are treatable and many are not. Diagnostic work-up to determine the etiology can be costly and invasive. Here we report the case of a man with slowly progressive spastic paraparesis. Using a multigene next-generation sequencing (NGS) panel, we identified a novel variant in the consensus splice site of the SPAST gene (exon 13, c.1536G>A, heterozygous), affecting codon 512 of the SPAST mRNA. The observed variant segregated with the disease in four tested family members. In this case, genetic confirmation obviated the need for additional testing such as MRI and lumbar puncture and helped the patient and his family understand his condition and prognosis. We conclude with a brief discussion of the SPG4/SPAST gene and the role of multigene panels in the diagnosis and management of hereditary spastic paraplegia.

A recurrent 16p12.1 microdeletion supports a two-hit model for severe developmental delay.

We report the identification of a recurrent, 520-kb 16p12.1 microdeletion associated with childhood developmental delay. The microdeletion was detected in 20 of 11,873 cases compared with 2 of 8,540 controls (P = 0.0009, OR = 7.2) and replicated in a second series of 22 of 9,254 cases compared with 6 of 6,299 controls (P = 0.028, OR = 2.5). Most deletions were inherited, with carrier parents likely to manifest neuropsychiatric phenotypes compared to non-carrier parents (P = 0.037, OR = 6). Probands were more likely to carry an additional large copy-number variant when compared to matched controls (10 of 42 cases, P = 5.7 x 10(-5), OR = 6.6). The clinical features of individuals with two mutations were distinct from and/or more severe than those of individuals carrying only the co-occurring mutation. Our data support a two-hit model in which the 16p12.1 microdeletion both predisposes to neuropsychiatric phenotypes as a single event and exacerbates neurodevelopmental phenotypes in association with other large deletions or duplications. Analysis of other microdeletions with variable expressivity indicates that this two-hit model might be more generally applicable to neuropsychiatric disease.

Initial transcribed sequence mutations specifically affect promoter escape properties.

Promoter escape efficiency of E. coli RNA polymerase is guided by both the core promoter and the initial transcribed sequence (ITS). Here, we quantitatively examined the escape properties of 43 random initial sequence variants of the phage T5 N25 promoter. The position for promoter escape on all N25-ITS variants occurred at the +15/+16 juncture, unlike the +11/+12 juncture for the wild type N25. These variants further exhibited a 25-fold difference in escape efficiency. ITS changes favoring promoter escape showed a compositional bias that is unrelated to nucleotide substrate binding affinity for the initial positions. Comparing all variants, the natural N25 promoter emerges as having evolved an ITS optimal for promoter escape, giving a high level of productive synthesis after undergoing the shortest abortive program. We supplemented GreB to transcription reactions to better understand abortive initiation and promoter escape in vivo. GreB supplementation elevated productive RNA synthesis 2-5-fold by altering the abortive RNA pattern, decreasing the abundance of the medium (6-10 nt) to long (11-15 nt) abortive RNAs without changing the levels of short (2-5 nt) and very long abortive RNAs (16-20 nt). The GreB-refractive nature of short abortive RNA production may reflect a minimum length requirement of 4-5 bp of the RNA-DNA hybrid for maintaining the stability of initial or backtracked complexes. That the very long abortive RNAs are unaffected by GreB suggests that they are unlikely to be products of polymerase backtracking. How the ITS might influence the course of early transcription is discussed within the structural context of an initial transcribing complex.