Auditory Development between 7 and 11 Years: An Event-Related Potential (ERP) Study

Background: There is considerable uncertainty about the time-course of central auditory maturation. On some indices, children appear to have adult-like competence by school age, whereas for other measures development follows a protracted course. Methodology: We studied auditory development using auditory event-related potentials (ERPs) elicited by tones in 105 children on two occasions two years apart. Just over half of the children were 7 years initially and 9 years at follow-up, whereas the remainder were 9 years initially and 11 years at follow-up. We used conventional analysis of peaks in the auditory ERP, independent component analysis, and time-frequency analysis. Principal Findings: We demonstrated maturational changes in the auditory ERP between 7 and 11 years, both using conventional peak measurements, and time-frequency analysis. The developmental trajectory was different for temporal vs. fronto-central electrode sites. Temporal electrode sites showed strong lateralisation of responses and no increase of low-frequency phase-resetting with age, whereas responses recorded from fronto-central electrode sites were not lateralised and showed progressive change with age. Fronto-central vs. temporal electrode sites also mapped onto independent components with differently oriented dipole sources in auditory cortex. A global measure of waveform shape proved to be the most effective method for distinguishing age bands. Conclusions/Significance: The results supported the idea that different cortical regions mature at different rates. The ICC measure is proposed as the best measure of 'auditory ERP age'

Engineering and functional analysis of mitotic kinases through chemical genetics

During mitosis, multiple protein kinases transform the cytoskeleton and chromosomes into new and highly dynamic structures that mediate the faithful transmission of genetic information and cell division. However, the large number and strong conservation of mammalian kinases in general pose significant obstacles to interrogating them with small molecules, due to the difficulty in identifying and validating those which are truly selective. To overcome this problem, a steric complementation strategy has been developed, in which a bulky “gatekeeper” residue within the active site of the kinase of interest is replaced with a smaller amino acid, such as glycine or alanine. The enlarged catalytic pocket can then be targeted in an allele-specific manner with bulky purine analogs. This strategy provides a general framework for dissecting kinase function with high selectivity, rapid kinetics, and reversibility. In this chapter we discuss the principles and techniques needed to implement this chemical genetic approach in mammalian cells

Decoding Polo-like kinase 1 signaling along the kinetochore–centromere axis

Protein kinase signaling along the kinetochore-centromere axis is crucial to assure mitotic fidelity, yet its spatial coordination is obscure. Here, we examined how pools of human Polo-like kinase 1 (Plk1) within this axis control signaling events to elicit mitotic functions. To do this, we restricted active Plk1 to discrete subcompartments within the kinetochore-centromere axis using chemical genetics and decoded functional and phosphoproteomic signatures of each. We observe distinct phosphoproteomic and functional roles, suggesting that Plk1 exists and functions in discrete pools along this axis. Deep within the centromere, Plk1 operates to assure proper chromosome alignment and segregation. Thus, Plk1 at the kinetochore is a conglomerate of an observable bulk pool coupled with additional functional pools below the threshold of microscopic detection/resolution. Although complex, this multiplicity of locales provides an opportunity to decouple functional and phosphoproteomic signatures for a comprehensive understanding of Plk1’s kinetochore functions

High-throughput decoding of antitrypanosomal drug efficacy and resistance.

The concept of disease-specific chemotherapy was developed a century ago. Dyes and arsenical compounds that displayed selectivity against trypanosomes were central to this work, and the drugs that emerged remain in use for treating human African trypanosomiasis (HAT). The importance of understanding the mechanisms underlying selective drug action and resistance for the development of improved HAT therapies has been recognized, but these mechanisms have remained largely unknown. Here we use all five current HAT drugs for genome-scale RNA interference target sequencing (RIT-seq) screens in Trypanosoma brucei, revealing the transporters, organelles, enzymes and metabolic pathways that function to facilitate antitrypanosomal drug action. RIT-seq profiling identifies both known drug importers and the only known pro-drug activator, and links more than fifty additional genes to drug action. A bloodstream stage-specific invariant surface glycoprotein (ISG75) family mediates suramin uptake, and the AP1 adaptin complex, lysosomal proteases and major lysosomal transmembrane protein, as well as spermidine and N-acetylglucosamine biosynthesis, all contribute to suramin action. Further screens link ubiquinone availability to nitro-drug action, plasma membrane P-type H(+)-ATPases to pentamidine action, and trypanothione and several putative kinases to melarsoprol action. We also demonstrate a major role for aquaglyceroporins in pentamidine and melarsoprol cross-resistance. These advances in our understanding of mechanisms of antitrypanosomal drug efficacy and resistance will aid the rational design of new therapies and help to combat drug resistance, and provide unprecedented molecular insight into the mode of action of antitrypanosomal drugs