The Parkinson’s disease kinase LRRK2 autophosphorylates its GTPase domain at multiple sites

Mutations in leucine-rich repeat kinase 2 (LRRK2) are a common cause of inherited Parkinson’s disease (PD). The protein is large and complex, but pathogenic mutations cluster in a region containing GTPase and kinase domains. LRRK2 can autophosphorylate in vitro within a dimer pair, although the significance of this reaction is unclear. Here, we mapped the sites of autophosphorylation within LRRK2 and found several potential phosphorylation sites within the GTPase domain. Using mass spectrometry, we found that Thr1343 is phosphorylated and, using kinase dead versions of LRRK2, show that this is an autophosphorylation site. However, we also find evidence for additional sites in the GTPase domain and in other regions of the protein suggesting that there may be multiple autophosphorylation sites within LRRK2. These data suggest that the kinase and GTPase activities of LRRK2 may exhibit complex autoregulatory interdependence.     

Synthesis and appetite suppressant activity of 1-aryloxy-2-substituted aminomethyltetrahydronaphthalenes as conformationally rigid analogues of fluoxetine

Several 1-aryloxy-2-substituted aminomethyltetrahydronaphthalenes (7-21) as conformationally rigid analogues of fluoxetine were synthesized and evaluated for their anorexigenic and antidepressant activities. For SAR studies the related acyclic analogues (22-27) were also prepared. Out of the 21 synthesized compounds, 10 compounds (9, 10, 11, 15, 16, 18, 21, 22, 23 and 27) exhibited significant anorexigenic activity (at 75 micromol/kg). Interestingly, all the compounds (7-20, 22-26) were devoid of antidepressant effect, except for compounds 21 and 27 in which the antidepressant activity was retained. Compound 16 emerged as the most active compound of the series with better anorexigenic activity (97.92%) compared to fluoxetine (76.25%) and even with a clinically used drug sibutramine, thus providing a new structural lead for appetite suppressants.     

Sarcomere lesion damage occurs mainly in slow fibers of reloaded rat adductor longus muscles

Sarcomerelesions were previously observed with reloading of rat adductor longusmuscles after spaceflight and hindlimb unloading (HU).Spaceflown rats displayed more lesioned fibers in the"slow-fiber" region, suggesting a damage-susceptible fiber type.Unloading induces fast myosin expression in some slow fibers,generating hybrid fibers. We examined whether lesion damage differedamong slow-, hybrid-, and fast-fiber types in HU-reloaded adductorlongus muscles. Temporal HU for 5, 8, 11, 14, and 17 days revealed that hybrid fiber percent, detected by antimyosin immunostaining, peaked at29 ± 12% by 14 days. A 14-day HU followed by 12-14 h ofvoluntary reloading was performed to induce lesions.² analysis showed that slowfibers were preferentially damaged, accounting for 92 ± 5% oflesioned fibers; hybrid and fast fibers accounted for 7 ± 4 and<0.5%, respectively. Atrophy did not explain differential lesiondamage across fiber types, as slow and hybrid fibers atrophied to asimilar extent. Because active myofiber contractions are requisite forlesion formation, selective recruitment of slow fibers most likelyexplains their damage susceptibility.

     

Effectiveness of VIA, Pap, and HPV DNA testing in a cervical cancer screening program in a peri-urban community in Andhra Pradesh, India

Background

While many studies have compared the efficacy of Pap cytology, visual inspection with acetic acid (VIA) and human papillomavirus (HPV) DNA assays for the detection cervical intraepithelial neoplasia and cancer, few have evaluated the program effectiveness.

Methods and Findings

A population-based sample of 5603 women from Medchal Mandal in Andhra Pradesh, India were invited to participate in a study comparing Pap cytology, VIA, and HPV DNA screening for the detection of CIN3+. Participation in primary screening and all subsequent follow-up visits was rigorously tracked. A 20% random sample of all women screened, in addition to all women with a positive screening test result underwent colposcopy with directed biopsy for final diagnosis. Sensitivity, specificity, positive and negative predictive values were adjusted for verification bias. HPV testing had a higher sensitivity (100%) and specificity (90.6%) compared to Pap cytology (sensitivity  =  78.2%; specificity = 86.0%) and VIA (sensitivity = 31.6%; specificity = 87.5%). Since 58% of the sample refused involvement and another 28% refused colposcopy or biopsy, we estimated that potentially 87.6% of the total underlying cases of CIN3 and cancer may have been missed due to program failures.

Conclusions

We conclude that despite our use of available resources, infrastructure, and guidelines for cervical cancer screening implementation in resource limited areas, community participation and non-compliance remain the major obstacles to successful reduction in cervical cancer mortality in this Indian population. HPV DNA testing was both more sensitive and specific than Pap cytology and VIA. The use of a less invasive and more user-friendly primary screening strategy (such as self-collected swabs for HPV DNA testing) may be required to achieve the coverage necessary for effective reduction in cervical cancer mortality.     

Evo-devo: Hydra raises its Noggin

Noggin, along with other secreted bone morphogenetic protein (BMP) inhibitors, plays a crucial role in neural induction and neural tube patterning as well as in somitogenesis, cardiac morphogenesis and formation of the skeleton in vertebrates. The BMP signalling pathway is one of the seven fundamental pathways that drive embryonic development and pattern formation in animals. Understanding its evolutionary origin and role in pattern formation is, therefore, important to evolutionary developmental biology (evo-devo). We have studied the evolutionary origin of BMP–Noggin antagonism in hydra, which is a powerful diploblastic model to study evolution of pattern-forming mechanisms because of the unusual cellular dynamics during its pattern formation and its remarkable ability to regenerate. We cloned and characterized the noggin gene from hydra and found it to exhibit considerable similarity with its orthologues at the amino acid level. Microinjection of hydra Noggin mRNA led to duplication of the dorsoventral axis in Xenopus embryos, demonstrating its functional conservation across the taxa. Our data, along with those of others, indicate that the evolutionarily conserved antagonism between BMP and its inhibitors predates bilateral divergence. This article reviews the various roles of Noggin in different organisms and some of our recent work on hydra Noggin in the context of evolution of developmental signalling pathways.     

Autonomic Cardiovascular Responses in Acclimatized Lowlanders on Prolonged Stay at High Altitude: A Longitudinal Follow Up Study

Acute exposure to hypobaric hypoxia at high altitude is reported to cause sympathetic dominance that may contribute to the pathophysiology of high altitude illnesses. The effect of prolonged stay at high altitude on autonomic functions, however, remains to be explored. Thus, the present study aimed at investigating the effect of high altitude on autonomic neural control of cardiovascular responses by monitoring heart rate variability (HRV) during chronic hypobaric hypoxia. Baseline electrocardiography (ECG) data was acquired from the volunteers at mean sea level (MSL) (<250 m) in Rajasthan. Following induction of the study population to high altitude (4500–4800 m) in Ladakh region, ECG data was acquired from the volunteers after 6 months (ALL 6) and 18 months of induction (ALL 18). Out of 159 volunteers who underwent complete investigation during acquisition of baseline data, we have only included the data of 104 volunteers who constantly stayed at high altitude for 18 months to complete the final follow up after 18 months. HRV parameters, physiological indices and biochemical changes in serum were investigated. Our results show sympathetic hyperactivation along with compromise in parasympathetic activity in ALL 6 and ALL 18 when compared to baseline data. Reduction of sympathetic activity and increased parasympathetic response was however observed in ALL 18 when compared to ALL 6. Our findings suggest that autonomic response is regulated by two distinct mechanisms in the ALL 6 and ALL 18. While the autonomic alterations in the ALL 6 group could be attributed to increased sympathetic activity resulting from increased plasma catecholamine concentration, the sympathetic activity in ALL 18 group is associated with increased concentration of serum coronary risk factors and elevated homocysteine. These findings have important clinical implications in assessment of susceptibility to cardio-vascular risks in acclimatized lowlanders staying for prolonged duration at high altitude.