Coping with Space Neophobia in Drosophila melanogaster: The Asymmetric Dynamics of Crossing a Doorway to the Untrodden

We discover and examine within a wide phylogenetic perspective spatial neophobia, avoidance of untrodden terrain, in fruit flies, in an experimental setup that reduces the gap between the field and the laboratory. In our setup, fruit flies use a natal fruit as their origin, freely exploring for days their surroundings, which consists of a mixture of trodden and untrodden terrain. The interface between trodden and untrodden is, however, reduced in our setup to a wide doorway, opened within a surrounding wall. Crossing this doorway, characterized by a sharp contrast interface between trodden and untrodden, generates a behavior whose dynamics betrays the flies'' space neophobia. The moment-by-moment dynamics of crossing is remarkably similar to that reported in mouse models of anxiety. This means that neophobic behavior is either homologous across arthropods and vertebrates or, not less interesting, convergent, whereby the same behavior is mediated in the two phyla by two completely different schemata.     

A multiplex biomarker approach for the diagnosis of transitional cell carcinoma from canine urine

Transitional cell carcinoma (TCC), the most common cancer of the urinary bladder in dogs, is usually diagnosed at an advanced disease stage with limited response to chemotherapy. Commercial screening tests lack specificity and current diagnostic procedures are invasive. A proof of concept pilot project for analyzing the canine urinary proteome as a noninvasive diagnostic tool for TCC identification was conducted. Urine was collected from 12 dogs in three cohorts (healthy, urinary tract infection, TCC) and analyzed using liquid chromatography tandem mass spectrometry. The presence of four proteins (macrophage capping protein, peroxiredoxin 5, heterogeneous nuclear ribonucleoproteins A2/B, and apolipoprotein A1) was confirmed via immunoblot. Of the total 379 proteins identified, 96 were unique to the TCC group. A statistical model, designed to evaluate the accuracy of this multiplex biomarker approach for diagnosis of TCC, predicted the presence of disease with 90% accuracy.     

Elg1, an alternative subunit of the RFC clamp loader,preferentially interacts with SUMOylated PCNA

Replication‐factor C (RFC) is a protein complex that loads the processivity clamp PCNA onto DNA. Elg1 is a conserved protein with homology to the largest subunit of RFC, but its function remained enigmatic. Here, we show that yeast Elg1 interacts physically and genetically with PCNA, in a manner that depends on PCNA modification, and exhibits preferential affinity for SUMOylated PCNA. This interaction is mediated by three small ubiquitin‐like modifier (SUMO)‐interacting motifs and a PCNA‐interacting protein box close to the N‐terminus of Elg1. These motifs are important for the ability of Elg1 to maintain genomic stability. SUMOylated PCNA is known to recruit the helicase Srs2, and in the absence of Elg1, Srs2 and SUMOylated PCNA accumulate on chromatin. Strains carrying mutations in both ELG1 and SRS2 exhibit a synthetic fitness defect that depends on PCNA modification. Our results underscore the importance of Elg1, Srs2 and SUMOylated PCNA in the maintenance of genomic stability.     

Cohesin Is Limiting for the Suppression of DNA Damage–Induced Recombination between Homologous Chromosomes

Double-strand break (DSB) repair through homologous recombination (HR) is an evolutionarily conserved process that is generally error-free. The risk to genome stability posed by nonallelic recombination or loss-of-heterozygosity could be reduced by confining HR to sister chromatids, thereby preventing recombination between homologous chromosomes. Here we show that the sister chromatid cohesion complex (cohesin) is a limiting factor in the control of DSB repair and genome stability and that it suppresses DNA damage–induced interactions between homologues. We developed a gene dosage system in tetraploid yeast to address limitations on various essential components in DSB repair and HR. Unlike RAD50 and RAD51, which play a direct role in HR, a 4-fold reduction in the number of essential MCD1 sister chromatid cohesion subunit genes affected survival of gamma-irradiated G₂/M cells. The decreased survival reflected a reduction in DSB repair. Importantly, HR between homologous chromosomes was strongly increased by ionizing radiation in G₂/M cells with a single copy of MCD1 or SMC3 even at radiation doses where survival was high and DSB repair was efficient. The increased recombination also extended to nonlethal doses of UV, which did not induce DSBs. The DNA damage–induced recombinants in G₂/M cells included crossovers. Thus, the cohesin complex has a dual role in protecting chromosome integrity: it promotes DSB repair and recombination between sister chromatids, and it suppresses damage-induced recombination between homologues. The effects of limited amounts of Mcd1and Smc3 indicate that small changes in cohesin levels may increase the risk of genome instability, which may lead to genetic diseases and cancer.     

Early molecular effects of ethanol during vertebrate embryogenesis

Fetal alcohol spectrum disorder (FASD) is the combination of developmental, morphological, and neurological defects that result from exposing human embryos to ethanol (EtOH). Numerous embryonic structures are affected, leading to a complex viable phenotype affecting among others, the anterior/posterior axis, head, and eye formation. Recent studies have provided evidence suggesting that EtOH teratogenesis is mediated in part through a reduction in retinoic acid (RA) levels, targeting mainly the embryonic organizer (Spemann's organizer) and its subsequent functions. EtOH-treated Xenopus embryos were subjected to an analysis of gene expression patterns. Analysis of organizer-specific genes revealed a transient delay in the invagination of gsc- and chordin-positive cells that eventually reach their normal rostro-caudal position. Dorsal midline genes show defects along the rostro-caudal axis, lacking either their rostral (Xbra and Xnot2) or caudal (FoxA4b and Shh) expression domains. Head-specific markers like Otx2, en2, and Shh show abnormal expression patterns. Otx2 exhibits a reduction in expression levels, while en2 becomes restricted along the dorsal/ventral axis. During neurula stages, Shh becomes up-regulated in the rostral region and it is expressed in an abnormal pattern. These results and histological analysis suggest the existence of malformations in the brain region including a lack of the normal fore brain ventricle. An increase in the size of both the prechordal plate and the notochord was observed, while the spinal cord is narrower. The reduction in head and eye size was accompanied by changes in the eye markers, Pax6 and Tbx3. Our results provide evidence for the early molecular changes induced by EtOH exposure during embryogenesis, and may explain some of the structural changes that are part of the EtOH teratogenic phenotype also in FASD individuals.     

RTVP-1 expression is regulated by SRF downstream of protein kinase C and contributes to the effect of SRF on glioma cell migration

Gliomas are characterized by increased infiltration into the surrounding normal brain tissue. We recently reported that RTVP-1 is highly expressed in gliomas and plays a role in the migration of these cells, however the regulation of RTVP-1 expression in these cells is not yet described. In this study we examined the role of PKC in the regulation of RTVP-1 expression and found that PMA and overexpression of PKCα and PKCε increased the expression of RTVP-1, whereas PKCδ exerted an opposite effect. Using the MatInspector software, we identified a SRF binding site on the RTVP-1 promoter. Chromatin immunoprecipitation (ChIP) assay revealed that SRF binds to the RTVP-1 promoter in U87 cells, and that this binding was significantly increased in response to serum addition. Moreover, silencing of SRF blocked the induction of RTVP-1 expression in response to serum. We found that overexpression of PKCα and PKCε increased the activity of the RTVP-1 promoter and the binding of SRF to the promoter. In contrast, overexpression of PKCδ blocked the increase in RTVP-1 expression in response to serum and the inhibitory effect of PKCδ was abrogated in cells expressing a SRFT160A mutant. SRF regulated the migration of glioma cells and its effect was partially mediated by RTVP-1. We conclude that RTVP-1 is a PKC-regulated gene and that this regulation is at least partly mediated by SRF. Moreover, RTVP-1 plays a role in the effect of SRF on glioma cell migration.     

Targeting telomerase-expressing cancer cells

The role of telomeres and telomerase as a target for cancer therapeutics is an area of continuing interest. This review is intended to provide an update on the field, pointing to areas in which our knowledge remains deficient and exploring the details of the most promising areas being advanced into clinical trials. Topics that will be covered include the role of dysfunctional telomeres in cellular aging and how replicative senescence provides an initial barrier to the emergence of immortalized cells, a hallmark of cancer. As an important translational theme, this review will consider possibilities for selectively targeting telomeres and telomerase to enhance cancer therapy. The role of telomerase as an immunotherapy, as a gene therapy approach using telomerase promoter driven oncolytic viruses and as a small oligonucleotide targeted therapy (Imetelstat) will be discussed.     

Histone deacetylase inhibitors for purging HIV-1 from the latent reservoir

A reservoir of latently infected memory CD4(+) T cells is believed to be the source of HIV-1 reemergence after discontinuation of antiretroviral therapy. HIV-1 eradication may depend on depletion of this reservoir. Integrated HIV-1 is inaccessible for expression, in part because of histone deacetylases (HDACs). One approach is to exploit the ability of HDAC inhibitors to induce HIV-1 expression from an integrated virus. With effective antiretroviral therapy, newly expressed HIV-1 is incapable of reinfecting naive cells. With HIV-1 expression, one assumes the infected cell dies and there is a progressive reduction in the size of the reservoir. The concept was tested using the HDAC inhibitor valproic acid. However, valproic acid is weak in inducing HIV-1 from latency in vitro. As such, clinical trials revealed a small or no effect on reducing the number of latently infected T cells in the peripheral blood. However, the new HDAC inhibitors vorinostat, belinostat and givinostat are more effective at targeting specific HDACs for HIV-1 expression than valproic acid. Here, we review studies on HDAC inhibitor-induced expression of latent HIV-1, with an emphasis on new and specific HDAC inhibitors. With increased potency for HIV-1 expression as well as safety and ease of oral administration, new HDAC inhibitors offer a unique opportunity to deplete the latent reservoir. An additional benefit is the antiinflammatory properties of HDAC inhibitors, including downregulation of HIV-1 coreceptor expression.     

Lower extremity fat mass is associated with insulin resistance in overweight and obese individuals: the CARDIA study

Lower extremity fat mass (LEFM) has been shown to be favorably associated with glucose metabolism. However, it is not clear whether this relationship is similar across varying levels of obesity. We hypothesized that lower amounts of LEFM is associated with higher insulin resistance (IR) and this association may vary according to weight status. Participants with available measures were examined from the Coronary Artery Risk Development in Young Adults study (CARDIA), a multi-center longitudinal study of the etiology of atherosclerosis in black and white men and women aged 38-50 years old in 2005-2006 (n = 1,579). The homeostasis model assessment of IR (HOMA(IR)) was calculated to estimate IR, regional adiposity was measured using dual energy X-ray absorptiometry (DXA), and weight status was defined according to BMI categories. Obese and overweight participants exhibited higher IR, total fat mass (FM), trunk FM (TFM), and LEFM compared to normal weight participants. After controlling for age, height, race, study center, education, smoking, and cardiorespiratory fitness (CRF), greater LEFM was significantly associated with higher IR only in normal weight men and women. Further adjustment for TFM revealed that lower LEFM was significantly associated with higher IR in overweight and obese men and women and the positive association in normal weight individuals was attenuated. These results suggest that excess adiposity in the lower extremities may attenuate the metabolic risk observed at a given level of abdominal adiposity in overweight and obese individuals. Weight status presents additional complexity since the metabolic influence of adipose tissue may not be homogenous across anatomic regions or level of obesity.