Effects of muscle model parameter dispersion and multi-loop segmental interaction on the neuromuscular system performance

The effects of parameter dispersion among motor units on the neuromuscular system performance as well as interaction between muscle segments and spinal cord mechanisms are investigated. Elementary components of the system are modeled to simulate with simple models their input-output characteristics. A leaky SS-IPFM encoder with a time-dependent threshold simulates the motor-neuron encoding characteristics. An amplitude and time dependent nonlinear model represent the motor unit mechanical output to neuronal input relationship. The dispersion of parameters in the components of the whole muscle control model is investigated in the open loop mode. It is shown that the dispersion of parameters in the multi-efferent channels converging on a common tendon provides a spatial filtration generating a smoother muscle force in addition to extending the linear dynamic range compared to a similar system having identical motor units. Muscle segmental interaction is investigated in this distributed model by closing the loop through a coupling matrix, representing afferent-motorneuron interaction on the spinal cord level. A diagonal matrix represents no segmental interaction and a uniform matrix represents a uniform interaction between segments through the muscle spindles and Golgi tendon feedback elements. The close loop simulation studied shows that (a). The type of segmental interaction has little effect on the overall system performance, i.e., range of linerity and stability, which is the result of having a muscle system with a large number of motor units. (b) There are only minor differences in results between the uniform and normal parameter distributions tested. (c) A loop gain of 4 divided by 8 in the distributed model can provide linearity through the full physiological force range. (d) Type of segmental interaction has significant effects on the individual segment. A uniform matrix provides a more stable segment due to the spatial filtration resulting from the segmental interaction, while the diagonal noninteracting matrix shows instabilities on the local segmental level despite global stability. The more realistic exponentially decaying spatial interaction matrix yields both global neuromuscular and local segmental stability with the same linear dynamic range generated with the uniform or diagonal matrices.     

Heritable Custom Genomic Modifications in Caenorhabditis elegans via a CRISPR–Cas9 System

We adapted the CRISPR–Cas9 system for template-mediated repair of targeted double-strand breaks via homologous recombination in Caenorhabditis elegans, enabling customized and efficient genome editing. This system can be used to create specific insertions, deletions, and base pair changes in the germline of C. elegans.     

The Genographic Project public participation mitochondrial DNA database

The Genographic Project is studying the genetic signatures of ancient human migrations and creating an open-source research database. It allows members of the public to participate in a real-time anthropological genetics study by submitting personal samples for analysis and donating the genetic results to the database. We report our experience from the first 18 months of public participation in the Genographic Project, during which we have created the largest standardized human mitochondrial DNA (mtDNA) database ever collected, comprising 78,590 genotypes. Here, we detail our genotyping and quality assurance protocols including direct sequencing of the mtDNA HVS-I, genotyping of 22 coding-region SNPs, and a series of computational quality checks based on phylogenetic principles. This database is very informative with respect to mtDNA phylogeny and mutational dynamics, and its size allows us to develop a nearest neighbor-based methodology for mtDNA haplogroup prediction based on HVS-I motifs that is superior to classic rule-based approaches. We make available to the scientific community and general public two new resources: a periodically updated database comprising all data donated by participants, and the nearest neighbor haplogroup prediction tool.     

Integrated Microfluidics for Protein Modification Discovery

Protein post-translational modifications mediate dynamic cellular processes with broad implications in human disease pathogenesis. There is a large demand for high-throughput technologies supporting post-translational modifications research, and both mass spectrometry and protein arrays have been successfully utilized for this purpose. Protein arrays override the major limitation of target protein abundance inherently associated with MS analysis. This technology, however, is typically restricted to pre-purified proteins spotted in a fixed composition on chips with limited life-time and functionality. In addition, the chips are expensive and designed for a single use, making complex experiments cost-prohibitive. Combining microfluidics with in situ protein expression from a cDNA microarray addressed these limitations. Based on this approach, we introduce a modular integrated microfluidic platform for multiple post-translational modifications analysis of freshly synthesized protein arrays (IMPA). The system''s potency, specificity and flexibility are demonstrated for tyrosine phosphorylation and ubiquitination in quasicellular environments. Unlimited by design and protein composition, and relying on minute amounts of biological material and cost-effective technology, this unique approach is applicable for a broad range of basic, biomedical and biomarker research.Protein post-translational modifications (PTMs)¹ vastly diversify eukaryotic proteomes and are integrated in essentially all cellular processes (1). Proteomic approaches, such as mass spectrometry (MS), have been instrumental in monitoring global molecular dynamics for research and clinical applications (2–5). However, even in this modern era, large-scale analyses of PTMs by MS is challenging because of the limited number of modified peptides derived from proteins that, by themselves, may not be abundant. Moreover, comprehensive PTM analysis by MS often requires significant amounts of biological material that may not be available. PTM analysis using protein arrays can overcome these limitations because of the equimolar amount of the arrayed proteins (6, 7). Large-scale protein arrays have been successfully integrated into PTM research (8, 9). However, this technology relies on pre-purified proteins that are arrayed on a surface and thus, incompatible with biochemically challenging proteins, let alone insoluble proteins. Moreover, the production of recombinant protein arrays is impractical in-house. Therefore, such arrays cannot be used fresh, and they are inherently limited to certain designs, protein compositions, and model organisms of high commercial value. To overcome the abovementioned limitations, we designed a modular integrated microfluidic platform for PTM analysis (IMPA).     

Coupling between Residues on S4 and S1 Defines the Voltage-Sensor Resting Conformation in NaChBac

The voltage sensor is a four-transmembrane helix bundle (S1-S4) that couples changes in membrane potential to conformational alterations in voltage-gated ion channels leading to pore opening and ion conductance. Although the structure of the voltage sensor in activated potassium channels is available, the conformation of the voltage sensor at rest is still obscure, limiting our understanding of the voltage-sensing mechanism. By employing a heterologously expressed Bacillus halodurans sodium channel (NaChBac), we defined constraints that affect the positioning and depolarization-induced outward motion of the S4 segment. We compared macroscopic currents mediated by NaChBac and mutants in which E43 on the S1 segment and the two outermost arginines (R1 and R2) on S4 were substituted. Neutralization of the negatively charged E43 (E43C) had a significant effect on channel gating. A double-mutant cycle analysis of E43 and R1 or R2 suggested changes in pairing during channel activation, implying that the interaction of E43 with R1 stabilizes the voltage sensor in its closed/available state, whereas interaction of E43 with R2 stabilizes the channel open/unavailable state. These constraints on S4 dynamics that define its stepwise movement upon channel activation and positioning at rest are novel, to the best of our knowledge, and compatible with the helical-screw and electrostatic models of S4 motion.     

The matrilineal ancestry of Ashkenazi Jewry: portrait of a recent founder event

Both the extent and location of the maternal ancestral deme from which the Ashkenazi Jewry arose remain obscure. Here, using complete sequences of the maternally inherited mitochondrial DNA (mtDNA), we show that close to one-half of Ashkenazi Jews, estimated at 8,000,000 people, can be traced back to only 4 women carrying distinct mtDNAs that are virtually absent in other populations, with the important exception of low frequencies among non-Ashkenazi Jews. We conclude that four founding mtDNAs, likely of Near Eastern ancestry, underwent major expansion(s) in Europe within the past millennium.     

The human cerebrospinal fluid metabolome

With continuing improvements in analytical technology and an increased interest in comprehensive metabolic profiling of biofluids and tissues, there is a growing need to develop comprehensive reference resources for certain clinically important biofluids, such as blood, urine and cerebrospinal fluid (CSF). As part of our effort to systematically characterize the human metabolome we have chosen to characterize CSF as the first biofluid to be intensively scrutinized. In doing so, we combined comprehensive NMR, gas chromatography-mass spectrometry (GC-MS) and liquid chromatography (LC) Fourier transform-mass spectrometry (FTMS) methods with computer-aided literature mining to identify and quantify essentially all of the metabolites that can be commonly detected (with today's technology) in the human CSF metabolome. Tables containing the compounds, concentrations, spectra, protocols and links to disease associations that we have found for the human CSF metabolome are freely available at http://www.csfmetabolome.ca.     

Counting the founders: the matrilineal genetic ancestry of the Jewish Diaspora

The history of the Jewish Diaspora dates back to the Assyrian and Babylonian conquests in the Levant, followed by complex demographic and migratory trajectories over the ensuing millennia which pose a serious challenge to unraveling population genetic patterns. Here we ask whether phylogenetic analysis, based on highly resolved mitochondrial DNA (mtDNA) phylogenies can discern among maternal ancestries of the Diaspora. Accordingly, 1,142 samples from 14 different non-Ashkenazi Jewish communities were analyzed. A list of complete mtDNA sequences was established for all variants present at high frequency in the communities studied, along with high-resolution genotyping of all samples. Unlike the previously reported pattern observed among Ashkenazi Jews, the numerically major portion of the non-Ashkenazi Jews, currently estimated at 5 million people and comprised of the Moroccan, Iraqi, Iranian and Iberian Exile Jewish communities showed no evidence for a narrow founder effect, which did however characterize the smaller and more remote Belmonte, Indian and the two Caucasus communities. The Indian and Ethiopian Jewish sample sets suggested local female introgression, while mtDNAs in all other communities studied belong to a well-characterized West Eurasian pool of maternal lineages. Absence of sub-Saharan African mtDNA lineages among the North African Jewish communities suggests negligible or low level of admixture with females of the host populations among whom the African haplogroup (Hg) L0-L3 sub-clades variants are common. In contrast, the North African and Iberian Exile Jewish communities show influence of putative Iberian admixture as documented by mtDNA Hg HV0 variants. These findings highlight striking differences in the demographic history of the widespread Jewish Diaspora.