Transduction of E2F-1 TAT fusion proteins represses expression of hTERT in primary ductal breast carcinoma cell lines

<p>Abstract</p> <p>Background</p> <p>Telomerase expression is detectable in 81–95% of breast carcinomas and may serve as a therapeutic target. The objective of this study was to investigate repression of telomerase activity in primary ductal breast cancer cells through transcriptional regulation of the catalytic subunit hTERT. We hypothesized that inhibition of telomerase expression could be achieved via Tat mediated protein transduction of the repressor protein E2F-1.</p> <p>Methods</p> <p>Protein purification techniques were refined to yield biologically active Tat fusion proteins (TFPs) capable of transducing the breast cancer cell lines HCC1937 and HCC1599. Cell lines were treated with wildtype E2F-1 (E2F-1/TatHA), mutant E2F-1 (E132/TatHA) and a control Tat peptide (TatHA) for 24 hours. Total RNA was isolated from treated cells, reverse transcribed and fold changes in gene expression for hTERT determined via real-time RT-qPCR.</p> <p>Results</p> <p>Significant repression of the catalytic subunit of telomerase (hTERT) was present in both HCC1937 and HCC1599 cells following treatment with E2F-1/TatHA. In HCC1937 cells, hTERT was repressed 3.5-fold by E2F-1/TatHA in comparison to E132/TatHA (p < 0.0012) and the TatHA peptide controls (p < 0.0024). In HCC1599 cells, hTERT was also repressed with E2F-1/TatHA treatment by 4.0-fold when compared to the E132/TatHA control (p < 0.0001). A slightly lower hTERT repression of 3.3-fold was observed with E2F-1/TatHA in the HCC1599 cells when compared to the TatHA control (p < 0.0001).</p> <p>Conclusion</p> <p>These results suggest that transduction of E2F-1/TatHA fusion proteins in vitro is an effective repressor of hTERT expression in the primary ductal breast cancer cell lines HCC1937 and HCC1599.</p

A Survey for Planetary Nebulae in M31 Globular Clusters

We report the results of an [O III] 5007 spectroscopic survey for planetary nebulae (PNe) located within the star clusters of M31. By examining R ~ 5000 spectra taken with the WIYN+Hydra spectrograph, we identify 3 PN candidates in a sample of 274 likely globular clusters, 2 candidates in objects which may be globular clusters, and 5 candidates in a set of 85 younger systems. The possible PNe are all faint, between ~2.5 and ~6.8 mag down the PN luminosity function, and, partly as a consequence of our selection criteria, have high excitation, with [O III] 5007 to H-beta ratios ranging from 2 to ~12. We discuss the individual candidates, their likelihood of cluster membership, and the possibility that they were formed via binary interactions within the clusters. Our data are consistent with the suggestion that PN formation within globular clusters correlates with binary encounter frequency, though, due to the small numbers and large uncertainties in the candidate list, this study does not provide sufficient evidence to confirm the hypothesis.Comment: Accepted for publication in the Astrophysical Journal. 54 pages, including 9 figures and 4 table

Signal peptide peptidases and gamma-secretase: Cousins of the same protease family?

Signal peptide peptidase (SPIP) is an unusual aspartyl protease, which mediates clearance of signal peptides by proteolysis within the endoplasmic reticulum (ER). Like presenilins, which provide the proteolytically active subunit of the,gamma-secretase complex, SPP contains a conserved GxGD motif in its C-terminal domain which is critical for its activity. While SPIP is known to be an aspartyl protease of the GxGD type, several presenilin homologues/SPP-like proteins (PSHs/ SPPL) of unknown function have been identified by database searches. In contrast to SPP and SPPL3, which are both restricted to the endoplasmic reticulum, SPPL2b is targeted through the secretory pathway to endosomes/lysosomes. As suggested by the differential subcellular localization of SPPL2b and SPPL3 distinct phenotypes were found upon antisense gripNA-mediated knockdown in zebrafish. spp and sppl3 knockdowns in zebrafish result in cell death within the central nervous system, whereas reduction of sppl2b expression causes erythrocyte accumulation in an enlarged caudal vein. Moreover, expression of D/A mutants of the putative C-terminal active sites of spp, sppl2, and spp13 produced phenocopies of the respective knockdown phenotypes. These data suggest that all investigated PSHs/SPPLs are members of the novel family of GxGD aspartyl proteases. More recently, it was shown that SPPL2b utilizes multiple intramembrane cleavages to liberate the TNF(x intracellular domain into the cytosol and to release the C-terminal counterpart into the lumen. These findings suggest common principles of intramembrane proteolysis by GxGD type aspartyl proteases. In this article,we will review the similarities of SPPs and gamma-secretase based on recent findings by us and others

The virtual peripheral nerve academy: education for the identification and treatment of peripheral nerve disorders

Millions of people around the globe suffer peripheral nerve injuries caused by trauma and medical disorders. However, medical school curricula are profoundly deficient in peripheral nerve education. This lack of knowledge within the healthcare profession may cause inadequate patient care. We developed the Virtual Peripheral Nerve Academy (VPNA) as a reusable virtual learning environment to provide medical students with detailed education on the peripheral nervous system (PNS). Students are introduced to the PNS through virtual 3D rendering of the human body, wherein they visualize individual nerves through dissection and observe normal motor and sensory function associated with each nerve. PNS structures that are absent from traditional texts are included in this visualization, ranging from the innervation of joints to the normal anatomic variation required for differential diagnosis of pain after an injury. Detailed modules on peripheral nerve disorders allow students to observe pathophysiological mechanisms, associated symptomatology, and appropriate treatments. Students are briefed on a patient clinical case, then interact with a patient avatar to learn the appropriate diagnostics, including physical exam maneuvers and electrodiagnostic testing. Interactive modules on peripheral nerve surgeries detail surgical techniques. The VPNA data and analytics dashboards allow medical students and course instructors to assess skill improvement and identify specific learning needs. The built-in learner management system and availability on both computer-based and virtual reality platforms facilitate integration into any existing medical school curricula. Ultimately, this immersive technology enables every medical student to learn about the peripheral nervous system and gain competency in treating real-life nerve pathologies

Spatiotemporal dynamics of the postnatal developing primate brain transcriptome.

Developmental changes in the temporal and spatial regulation of gene expression drive the emergence of normal mature brain function, while disruptions in these processes underlie many neurodevelopmental abnormalities. To solidify our foundational knowledge of such changes in a primate brain with an extended period of postnatal maturation like in human, we investigated the whole-genome transcriptional profiles of rhesus monkey brains from birth to adulthood. We found that gene expression dynamics are largest from birth through infancy, after which gene expression profiles transition to a relatively stable state by young adulthood. Biological pathway enrichment analysis revealed that genes more highly expressed at birth are associated with cell adhesion and neuron differentiation, while genes more highly expressed in juveniles and adults are associated with cell death. Neocortex showed significantly greater differential expression over time than subcortical structures, and this trend likely reflects the protracted postnatal development of the cortex. Using network analysis, we identified 27 co-expression modules containing genes with highly correlated expression patterns that are associated with specific brain regions, ages or both. In particular, one module with high expression in neonatal cortex and striatum that decreases during infancy and juvenile development was significantly enriched for autism spectrum disorder (ASD)-related genes. This network was enriched for genes associated with axon guidance and interneuron differentiation, consistent with a disruption in the formation of functional cortical circuitry in ASD

Strategic and Interactive Writing Instruction: An efficacy study in grades 3-5

A quasi-experimental study was conducted to examine the impact of Strategic and Interactive Writing Instruction on 3rd-5th grade deaf and hard of hearing students’ writing and language compared to a business-as-usual condition (N=63). A total of 18 hours of instruction was provided for each of two types of writing—personal narrative (known as recount) and persuasive. Writing samples, collected prior to instruction and after, were scored for writing traits, language accuracy and complexity. Data were analyzed using a two-level, mixed-effects regression. Results show the treatment to be effective for recount and persuasive writing traits, and recount language variables, with effect sizes ranging from 0.46 to 2.01. Treatment effects were also substantial for persuasive writing language outcomes (0.38 to 1.06), although not all were statistically significant at the 0.05 level. The findings suggest the importance of apprenticeship in writing and consideration for the specific language needs of students with hearing loss

Resin and Magnetic Nanoparticle-Based Free Radical Probes for Glycan Capture, Isolation, and Structural Characterization

By combining the merits of solid supports and free radical activated glycan sequencing (FRAGS) reagents, we develop a multifunctional solid-supported free radical probe (SS-FRAGS) that enables glycan enrichment and characterization. SS-FRAGS comprises a solid support, free radical precursor, disulfide bond, pyridyl, and hydrazine moieties. Thio-activated resin and magnetic nanoparticles (MNPs) are chosen as the solid support to selectively capture free glycans via the hydrazine moiety, allowing for their enrichment and isolation. The disulfide bond acts as a temporary covalent linkage between the solid support and the captured glycan, allowing the release of glycans via the cleavage of the disulfide bond by dithiothreitol. The basic pyridyl functional group provides a site for the formation of a fixed charge, enabling detection by mass spectrometry and avoiding glycan rearrangement during collisional activation. The free radical precursor generates a nascent free radical upon collisional activation and thus simultaneously induces systematic and predictable fragmentation for glycan structure elucidation. A radical-driven glycan deconstruction diagram (R-DECON) is developed to visually summarize the MS² results and thus allow for the assembly of the glycan skeleton, making the differentiation of isobaric glycan isomers unambiguous. For application to a real-world sample, we demonstrate the efficacy of the SS-FRAGS by analyzing glycan structures enzymatically cleaved from RNase-B

Phytoplankton blooms weakly influence the cloud forming ability of sea spray aerosol

After many field studies, the establishment of connections between marine microbiological processes, sea spray aerosol (SSA) composition, and cloud condensation nuclei (CCN) has remained an elusive challenge. In this study, we induced algae blooms to probe how complex changes in seawater composition impact the ability of nascent SSA to act as CCN, quantified by using the apparent hygroscopicity parameter (κapp). Throughout all blooms, κapp ranged between 0.7 and 1.4 (average 0.95 ± 0.15), consistent with laboratory investigations using algae‐produced organic matter, but differing from climate model parameterizations and in situ SSA generation studies. The size distribution of nascent SSA dictates that changes in κapp associated with biological processing induce less than 3% change in expected CCN concentrations for typical marine cloud supersaturations. The insignificant effect of hygroscopicity on CCN concentrations suggests that the SSA production flux and/or secondary aerosol chemistry may be more important factors linking ocean biogeochemistry and marine clouds.Key PointsChanges in seawater and sea spray composition did not strongly affect expected CCN concentrationsBlooms may impact clouds more strongly through changes in aerosol flux or secondary chemistryModel parameterizations likely overestimate changes in cloud nuclei due to primary marine organicsPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/134444/1/grl54978_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/134444/2/grl54978-sup-0001-supinfo.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/134444/3/grl54978.pd