A novel cell immunoassay to measure survival of motor neurons protein in blood cells

BACKGROUND: The motor neuron degenerative disease spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality and is caused by mutations in the survival of motor neurons (SMN) gene that reduce the expression levels of the SMN protein. A major goal of current therapeutic approaches is to increase SMN levels in SMA patients. The purpose of this study was to develop a reliable assay to measure SMN protein levels from peripheral blood samples. METHODS: We developed a novel cell immunoassay to quantitatively measure SMN levels from peripheral blood mononuclear cells (PBMCs) using a single anti-SMN antibody. RESULTS: SMN levels determined by the cell immunoassay are comparable to levels determined by Western blot, but in contrast, the immunoassay does not involve cell lysis, requires a small amount of patient material, and can be done on a large number of samples simultaneously. SMN levels from PBMCs are not influenced by cell type heterogeneity. CONCLUSION: SMN levels measured from total PBMCs provide an important snapshot of SMN protein expression, which should be a useful aid in SMA diagnosis, and a surrogate marker of efficacy of treatment in SMA clinical trials

Exome Sequencing Identifies a Novel TRPV4 Mutation in a CMT2C Family

Molecular and Cellular Biolog

Improvement of Neuromuscular Synaptic Phenotypes without Enhanced Survival and Motor Function in Severe Spinal Muscular Atrophy Mice Selectively Rescued in Motor Neurons

In the inherited childhood neuromuscular disease spinal muscular atrophy (SMA), lower motor neuron death and severe muscle weakness result from the reduction of the ubiquitously expressed protein survival of motor neuron (SMN). Although SMA mice recapitulate many features of the human disease, it has remained unclear if their short lifespan and motor weakness are primarily due to cell-autonomous defects in motor neurons. Using Hb9(Cre) as a driver, we selectively raised SMN expression in motor neurons in conditional SMAΔ7 mice. Unlike a previous study that used choline acetyltransferase (ChAT(Cre+) ) as a driver on the same mice, and another report that used Hb9(Cre) as a driver on a different line of conditional SMA mice, we found no improvement in survival, weight, motor behavior and presynaptic neurofilament accumulation. However, like in ChAT(Cre+) mice, we detected rescue of endplate size and mitigation of neuromuscular junction (NMJ) denervation status. The rescue of endplate size occurred in the absence of an increase in myofiber size, suggesting endplate size is determined by the motor neuron in these animals. Real time-PCR showed that the expression of spinal cord SMN transcript was sharply reduced in Hb9(Cre+) SMA mice relative to ChAT(Cre+) SMA mice. This suggests that our lack of overall phenotypic improvement is most likely due to an unexpectedly poor recombination efficiency driven by Hb9(Cre) . Nonetheless, the low levels of SMN were sufficient to rescue two NMJ structural parameters indicating that these motor neuron cell autonomous phenotypes are very sensitive to changes in motoneuronal SMN levels. Our results directly suggest that even those therapeutic interventions with very modest effects in raising SMN in motor neurons may provide mitigation of neuromuscular phenotypes in SMA patients

A motor neuron disease–associated mutation in p150Glued perturbs dynactin function and induces protein aggregation

The microtubule motor cytoplasmic dynein and its activator dynactin drive vesicular transport and mitotic spindle organization. Dynactin is ubiquitously expressed in eukaryotes, but a G59S mutation in the p150Glued subunit of dynactin results in the specific degeneration of motor neurons. This mutation in the conserved cytoskeleton-associated protein, glycine-rich (CAP-Gly) domain lowers the affinity of p150Glued for microtubules and EB1. Cell lines from patients are morphologically normal but show delayed recovery after nocodazole treatment, consistent with a subtle disruption of dynein/dynactin function. The G59S mutation disrupts the folding of the CAP-Gly domain, resulting in aggregation of the p150Glued protein both in vitro and in vivo, which is accompanied by an increase in cell death in a motor neuron cell line. Overexpression of the chaperone Hsp70 inhibits aggregate formation and prevents cell death. These data support a model in which a point mutation in p150Glued causes both loss of dynein/dynactin function and gain of toxic function, which together lead to motor neuron cell death

Dominant mutations in the cation channel gene transient receptor potential vanilloid 4 cause an unusual spectrum of neuropathies

Hereditary neuropathies form a heterogeneous group of disorders for which over 40 causal genes have been identified to date. Recently, dominant mutations in the transient receptor potential vanilloid 4 gene were found to be associated with three distinct neuromuscular phenotypes: hereditary motor and sensory neuropathy 2C, scapuloperoneal spinal muscular atrophy and congenital distal spinal muscular atrophy. Transient receptor potential vanilloid 4 encodes a cation channel previously implicated in several types of dominantly inherited bone dysplasia syndromes. We performed DNA sequencing of the coding regions of transient receptor potential vanilloid 4 in a cohort of 145 patients with various types of hereditary neuropathy and identified five different heterozygous missense mutations in eight unrelated families. One mutation arose de novo in an isolated patient, and the remainder segregated in families. Two of the mutations were recurrent in unrelated families. Four mutations in transient receptor potential vanilloid 4 targeted conserved arginine residues in the ankyrin repeat domain, which is believed to be important in protein-protein interactions. Striking phenotypic variability between and within families was observed. The majority of patients displayed a predominantly, or pure, motor neuropathy with axonal characteristics observed on electrophysiological testing. The age of onset varied widely, ranging from congenital to late adulthood onset. Various combinations of additional features were present in most patients including vocal fold paralysis, scapular weakness, contractures and hearing loss. We identified six asymptomatic mutation carriers, indicating reduced penetrance of the transient receptor potential vanilloid 4 defects. This finding is relatively unusual in the context of hereditary neuropathies and has important implications for diagnostic testing and genetic counsellin

The James Webb Space Telescope Mission

Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least $4m$. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the $6.5m$ James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 years, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space Telescope Overview, 29 pages, 4 figure