Dynamics of Dynamin during Clathrin Mediated Endocytosis in PC12 Cells

Members of the dynamin super-family of GTPases are involved in disparate cellular pathways. Dynamin1 and dynamin2 have been implicated in clathrin-mediated endocytosis. While some models suggest that dynamin functions specifically at the point of vesicle fission, evidence also exists for a role prior to fission during vesicle formation and it is unknown if there is a role for dynamin after vesicle fission. Although dynamin2 is ubiquitously expressed, dynamin1 is restricted to the nervous system. These two structurally similar endocytic accessory proteins have not been studied in cells that endogenously express both.The present study quantitatively assesses the dynamics of dynamin1 and dynamin2 during clathrin-mediated endocytosis in PC12 cells, which endogenously express both proteins. Both dynamin isoforms co-localized with clathrin and showed sharp increases in fluorescence intensity immediately prior to internalization of the nascent clathrin-coated vesicle. The fluorescence intensity of both proteins then decreased with two time constants. The slower time constant closely matched the time constant for the decrease of clathrin intensity and likely represents vesicle movement away from the membrane. The faster rate may reflect release of dynamin at the neck of nascent vesicle following GTP hydrolysis.This study analyses the role of dynamin in clathrin-mediated endocytosis in a model for cellular neuroscience and these results may provide direct evidence for the existence of two populations of dynamin associated with nascent clathrin-coated vesicles

Genomic Relationships, Novel Loci, and Pleiotropic Mechanisms across Eight Psychiatric Disorders

Genetic influences on psychiatric disorders transcend diagnostic boundaries, suggesting substantial pleiotropy of contributing loci. However, the nature and mechanisms of these pleiotropic effects remain unclear. We performed analyses of 232,964 cases and 494,162 controls from genome-wide studies of anorexia nervosa, attention-deficit/hyper-activity disorder, autism spectrum disorder, bipolar disorder, major depression, obsessive-compulsive disorder, schizophrenia, and Tourette syndrome. Genetic correlation analyses revealed a meaningful structure within the eight disorders, identifying three groups of inter-related disorders. Meta-analysis across these eight disorders detected 109 loci associated with at least two psychiatric disorders, including 23 loci with pleiotropic effects on four or more disorders and 11 loci with antagonistic effects on multiple disorders. The pleiotropic loci are located within genes that show heightened expression in the brain throughout the lifespan, beginning prenatally in the second trimester, and play prominent roles in neurodevelopmental processes. These findings have important implications for psychiatric nosology, drug development, and risk prediction.Peer reviewe

Barriers to acceptance of the human papillomavirus prophylactic vaccine

Abstract Human papillomavirus (HPV) is the most common sexually transmitted infection in the United States. In June 2006, the US Food and Drug Administration approved Gardasil, a quadrivalent HPV 6, 11, 16 and 18 recombinant vaccine. Indicated for young girls and women aged 9 to 26, Gardasil is the first vaccine approved to prevent transmission of HPV types 16 and 18. In order to move toward universal acceptance of the HPV vaccine by the general population, both mandated HPV vaccination and HPV vaccination-specific issues must be addressed. Identifying and understanding factors associated with the acceptance of the HPV vaccine has been and will continue to be important so physicians can assist parents and adolescents in their decision to refuse or accept the vaccine. Despite the potential social and cultural barriers to a universal HPV vaccination program, numerous studies have indicated a willingness among many women to accept the vaccine for their daughters. It is clear that education will play an important role in the implementation of such a vaccination program

Increase in dynamin2 intensity at sites of clathrin-mediated endocytosis in PC12 cells.

<p>PC12 cells co-transfected with clathrin-dsRed (A) and dynamin2-EGFP (B) were imaged by live-cell TIR-FM. Frames with time-stamps represent magnifications of regions identified directly to the left. Quantification of the spot marker by the arrow (C) shows burst of dynamin2 fluorescence just prior to endocytosis. Additionally, clathrin fluorescence remains roughly constant prior to internalization. Quantification of 15 such events is shown is (D).</p

Increase in dynamin1 intensity at sites of clathrin-mediated endocytosis in PC12 cells.

<p>PC12 cells co-transfected with clathrin-dsRed (A) and dynamin1-EGFP (B) were imaged by live-cell TIR-FM. Frames with time-stamps represent magnifications of regions identified directly to the left. Quantification of the spot marker by the arrow (C) shows burst of dynamin1 fluorescence just prior to endocytosis. Additionally, clathrin fluorescence remains roughly constant prior to internalization. Quantification of 12 such events is shown is (D).</p

Co-localization of clathrin and dynamin1 in PC12 cells.

<p>PC12 cells co-transfected with clathrin-dsRed (A,C) and dynamin1-EGFP (B,D) were imaged by TIR-FM. Regions drawn around clathrin spots (A) were transferred to dynamin1 images (red circles in D). Regions drawn around dynamin1 spots (B) were transferred to clathrin images (green circles in C). 250 spots per group were evaluated to determine if clathrin and dynamin1 co-localize (E). Dynamin1 at sites of clathrin spots is significantly higher than dynamin1 outside clathrin spots. Clathrin corresponding to dynamin1 spots is significantly higher than dynamin1 outside clathrin spots.</p

Different phases of dynamin intensity decrease at sites of clathrin-mediated endocytosis in PC12 cells.

<p>PC12 cells co-transfected with clathrin-dsRed and dynamin1-EGFP (A) or dynamin2-EGFP (B) were imaged by live-cell TIR-FM. Decreases in dynamin fluorescence were aligned so that the peak was set to time 0, and clathrin and Dynamin fluorescence at time 0 were normalized to 1. (B and C) demonstrate the decreases in clathrin and dynamin1 or dynamin2, at the sites of 30 or 28 disappearing clathrin spots, respectively. In each case, the decrease in clathrin intensity fits a single exponential, while both dynamin1 and dynamin2 fit double exponentials (all r² values >0.98).</p

Willingness of pregnant women to vaccinate themselves and their newborns with the HPV vaccine

To evaluate the willingness of pregnant women to accept the HPV vaccine for their newborns as well as themselves. An 18-item questionnaire was distributed to antepartum women. Demographic data about the respondent's current pregnancy and her knowledge of HPV and the HPV vaccine was collected. Information about the respondent's HPV and HPV vaccine status as well as her acceptance of the vaccine for herself during pregnancy and her newborn son and/or daughter after delivery was also collected. Three hundred surveys were completed and available for review. Only 6 respondents (2%) had received the HPV vaccine. Despite the small group of patients who had previously been vaccinated, 112 respondents (37.3%) stated that they would receive the HPV vaccine during pregnancy if recommended by an obstetrician. 99 respondents (33%) stated that they would vaccinate their newborn female infant and 83 (27.7%) stated would vaccinate their male infants. Providing the HPV vaccine to pregnant women as well as newborns could be an important way to increase the patient population who is protected against HPV. A percentage of pregnant women are willing to accept the vaccine for themselves and their newborns

Minimally Invasive Staging of Endometrial Cancer Is Feasible and Safe in Elderly Women

To compare the surgical outcome of elderly and younger patients undergoing laparoscopic or robotic surgical staging of endometrial cancer. Retrospective analysis (Canadian Task Force classification II-2). University-affiliated hospital. One hundred twenty-nine patients comprised the study group. Sixty patients were aged 65 years or older (elderly group), and 69 patients were younger than 65 years (younger group). Abdominal, laparoscopic, or robotic hysterectomy. Among the 109 patients who underwent laparoscopic or robotic staging, there were no differences in estimated blood loss, lymph node count, surgical time, complications, rate of blood transfusion, conversion to laparotomy, and mean postoperative stay between elderly and younger patients. Minimally invasive surgical staging for endometrial cancer is both feasible and safe in the elderly population and offers similar outcomes as in younger patients