Prospects for a Statistical Theory of LC/TOFMS Data

The critical importance of employing sound statistical arguments when seeking to draw inferences from inexact measurements is well-established throughout the sciences. Yet fundamental statistical methods such as hypothesis testing can currently be applied to only a small subset of the data analytical problems encountered in LC/MS experiments. The means of inference that are more generally employed are based on a variety of heuristic techniques and a largely qualitative understanding of their behavior. In this article, we attempt to move towards a more formalized approach to the analysis of LC/TOFMS data by establishing some of the core concepts required for a detailed mathematical description of the data. Using arguments that are based on the fundamental workings of the instrument, we derive and validate a probability distribution that approximates that of the empirically obtained data and on the basis of which formal statistical tests can be constructed. Unlike many existing statistical models for MS data, the one presented here aims for rigor rather than generality. Consequently, the model is closely tailored to a particular type of TOF mass spectrometer although the general approach carries over to other instrument designs. Looking ahead, we argue that further improvements in our ability to characterize the data mathematically could enable us to address a wide range of data analytical problems in a statistically rigorous manner

Characterization of the role of the Rab GTPase-activating protein AS160 in insulin-regulated GLUT4 trafficking

Insulin stimulates the translocation of the glucose transporter GLUT4 from intracellular vesicles to the plasma membrane. In the present study we have conducted a comprehensive proteomic analysis of affinity-purified GLUT4 vesicles from 3T3-L1 adipocytes to discover potential regulators of GLUT4 trafficking. In addition to previously identified components of GLUT4 storage vesicles including the insulin-regulated aminopeptidase insulin-regulated aminopeptidase and the vesicle soluble N-ethylmaleimide factor attachment protein (v-SNARE) VAMP2, we have identified three new Rab proteins, Rab10, Rab11, and Rab14, on GLUT4 vesicles. We have also found that the putative Rab GTPase-activating protein AS160 (Akt substrate of 160 kDa) is associated with GLUT4 vesicles in the basal state and dissociates in response to insulin. This association is likely to be mediated by the cytosolic tail of insulin-regulated aminopeptidase, which interacted both in vitro and in vivo with AS160. Consistent with an inhibitory role of AS160 in the basal state, reduced expression of AS160 in adipocytes using short hairpin RNA increased plasma membrane levels of GLUT4 in an insulin-independent manner. These findings support an important role for AS160 in the insulin regulated trafficking of GLUT4