Faculty Bibliography

The advent of new technologies for the imaging of living cells has made it possible to determine the properties of transcription, the kinetics of polymerase movement, the association of transcription factors, and the progression of the polymerase on the gene. We report here the current state of the field and the progress necessary to achieve a more complete understanding of the various steps in transcription. Our Consortium is dedicated to developing and implementing the technology to further this understanding.

Protein misfolding in the endoplasmic reticulum (ER) activates a set of intracellular signaling pathways, collectively termed the Unfolded Protein Response (UPR). UPR signaling promotes cell survival by reducing misfolded protein levels. If homeostasis cannot be restored, UPR signaling promotes cell death. The molecular basis for the switch between prosurvival and proapoptotic UPR function is poorly understood. The ER-resident proteins, PERK and IRE1, control two key UPR signaling pathways. Protein misfolding concomitantly activates PERK and IRE1 and has clouded insight into their contributions toward life or death cell fates. Here, we employed chemical-genetic strategies to activate individually PERK or IRE1 uncoupled from protein misfolding. We found that sustained PERK signaling impaired cell proliferation and promoted apoptosis. By contrast, equivalent durations of IRE1 signaling enhanced cell proliferation without promoting cell death. These results demonstrate that extended PERK and IRE1 signaling have opposite effects on cell viability. Differential activation of PERK and IRE1 may determine life or death decisions after ER protein misfolding

Neuronal ELAV (nELAV) proteins are RNA-binding proteins which play a physiological role in controlling gene expression in memory formation, and their alteration may contribute to cognitive impairment associated with neurodegenerative pathologies such as Alzheimer's disease (AD). Indeed, we found that the content of nELAV proteins is significantly decreased along with clinical dementia progression in the hippocampi of AD brains, where it inversely correlates with the amount of amyloid-beta (Abeta). To check the direct influence of Abeta on nELAV, we performed in vitro experiments using human SH-SY5Y cells, finding that Abeta(1-42) specifically determines nELAV proteins reduction. Since ADAM10 mRNA has the predicted sequences targeted by nELAV, we investigated whether Abeta, through nELAV proteins, could originate a vicious circle affecting amyloid-beta protein precursor (AbetaPP) processing. Immunoprecipitation experiments showed that indeed nELAV proteins bind to ADAM10 mRNA and that this binding is disrupted by Abeta(1-42) exposure, resulting in a decreased ADAM10 protein expression. ADAM10 protein diminution was also found in AD hippocampi. These data show for the first time the involvement of nELAV in AD pathology and suggest that their alteration may affect genes implicated in AbetaPP processing

The progressive ankylosis gene (ank) is a transmembrane protein that transports intracellular pyrophosphate to the extracellular milieu. Human mutations of ank lead to craniometaphyseal dysplasia, a disease which is characterized by the overgrowth of craniofacial bones and osteopenia in long bones, suggesting that ANK plays a regulatory role in osteoblast differentiation. To determine the role of ANK in osteoblast differentiation, we suppressed ANK expression in the osteoblastic MC3T3-E1 cell line using siRNA and determined the expression of osteoblastic marker genes and the transcription factors osterix and runx2. In addition, we determined the osteoblastic differentiation of bone marrow stromal cells isolated from the bone marrow of ank/ank mice, which express a truncated, nonfunctional ANK protein, or wild-type littermates. Suppression of ANK expression in MC3T3-E1 cells led to a decrease in bone marker gene expression, including alkaline phosphatase, bone sialoprotein, osteocalcin and type I collagen. In addition, osterix gene expression was decreased in ANK expression-suppressed MC3T3 cells, whereas runx2 expression was increased. Bone marrow stromal cells isolated from ank/ank mice cultured in the presence of ascorbate-2-phosphate for up to 35 days showed markedly reduced mineralization compared to the mineralization of bone marrow stromal cells isolated from wild-type littermates. In conclusion, these findings suggest that ANK is a positive regulator of differentiation events towards a mature osteoblastic phenotype

A strong link is indicated between cardiovascular disease (CVD) and risk for developing Alzheimer's disease (AD), which may be exacerbated by the major AD genetic risk factor apolipoprotein Eepsilon4 (APOEepsilon4). Since subjective memory complaint (SMC) may potentially be an early indicator for cognitive decline, we examined CVD risk factors in a cohort of SMC. As amyloid-beta (Abeta) is considered to play a central role in AD, we hypothesized that the CVD risk profile (increased LDL, reduced HDL, and increased body fat) would be associated with plasma Abeta levels. We explored this in 198 individuals with and without SMC (average age = 63 years). Correlations between Abeta40 and HDL were observed, which were stronger in non-APOEepsilon4 carriers (rho = -0.315, p < 0.001) and in SMC (rho = -0.322, p = 0.01). There was no relationship between percentage body fat and Abeta40 in this cohort. Age and HDL remained predictive for plasma Abeta40 using multivariate regression analysis. We report a novel negative association between HDL and Abeta, which if demonstrated to be causal has implications for the development of lifestyle interventions and/or novel therapeutics. The relationship between HDL and Abeta and the potential significance of such an association needs to be validated in a larger longitudinal study.

The mammalian skull vault consists mainly of 5 flat bones, the paired frontals and parietals, and the unpaired interparietal. All of these bones are formed by intramembranous ossification within a layer of mesenchyme, the skeletogenic membrane, located between the dermal mesenchyme and the meninges surrounding the brain. While the frontal bones are of neural crest in origin, the parietal bones arise from mesoderm. The present study is a characterization of frontal and parietal bones at their molecular level, aiming to highlight distinct differences between the neural crest-derived frontal and mesodermal-derived parietal bone. We performed a detailed comparative gene expression profile of FGF ligands and their receptors known to play crucial role in skeletogenesis. This analysis revealed that a differential expression pattern of the major FGF osteogenic molecules and their receptors exists between the neural crest-derived frontal bone and the paraxial mesoderm-derived parietal bone. Particularly, the expression of ligands such as Fgf-2, Fgf-9 and Fgf-18 was upregulated in frontal bone on embryonic day 17.5, postnatal day 1 and postnatal day 60 mice. Frontal bone also elaborated higher levels of Fgf receptor 1, 2 and 3 transcripts versus parietal bone. Taken together, these data suggest that the frontal bone is a domain with higher FGF-signaling competence than parietal bone.

Invasive aspergillosis remains difficult to diagnose despite advances in imaging and antigen-based serological testing. To overcome this problem, nucleic acid (NA)-based amplification assays were introduced to identify infecting pathogens. Unfortunately, the reliability of such assays to detect Aspergillus spp. has met with mixed success. A new generation of NA platforms are emerging, which greatly improve our ability to detect Aspergillus-specific DNA and RNA from respiratory and blood samples. These platforms can accurately detect a single genome, and the emergence of pan-fungal and pan-Aspergillus probes offer promise for broader detection. PCR remains the most important platform, especially when coupled with real-time probes. It is multiplex friendly and can distinguish between closely related target sequences. Nucleic acid sequence-based amplification (NASBA) is an RNA-directed isothermal transcription-based amplification platform, which is more robust than PCR resulting in a 10(14)-fold amplification. RNA-based detection facilitates more target options and can be used to assess cell viability. Both DNA and RNA amplification platforms take advantage of allele-specific properties of probes, which are valuable for assessing drug resistance markers. Finally, as new molecular diagnostic platforms mature, their role may expand to include early monitoring of therapy.

Sex determination mechanisms are thought to evolve rapidly and show little conservation among different animal species. For example, the critical gene on the Y chromosome, SRY, that determines sex in most mammals, is not found in other animals. However, a related Sox domain transcription factor, SOX9, is also required for testis development in mammals and exhibits male-specific gonad expression in other vertebrate species. Previously, we found that the Drosophila orthologue of SOX9, Sox100B, is expressed male-specifically during gonad development. We now investigate the function of Sox100B and find, strikingly, that Sox100B is essential for testis development in Drosophila. In Sox100B mutants, the adult testis is severely reduced and fails to interact with other parts of the reproductive tract, which are themselves unaffected. While a testis initially forms in Sox100B mutants, it fails to undergo proper morphogenesis during pupal stages, likely due to defects in the pigment cells. In contrast, no substantive defects are observed in ovary development in Sox100B mutant females. Thus, as is observed in mammals, a Sox9 homolog is essential for sex-specific gonad development in Drosophila, suggesting that the molecular mechanisms regulating sexually dimorphic gonad development may be more conserved than previously suspected.

Reports an error in 'Cerebrospinal fluid monoaminergic metabolites in wild Papio anubis and P. hamadryas are concordant with taxon-specific behavioral ontogeny' by Clifford J. Jolly, Jane E. Phillips-Conroy, Jay R. Kaplan and J. John Mann (International Journal of Primatology, 2008[Dec], Vol 29[6], 1549-1566). Due to a production error, the author corrections were not incorporated into the published version of the manuscript. 1. Improved versions of Figs. 1 and 2 are printed with their captions: Fig. 1 Fig. 2 2. Table 5, missing from the published version, is given on this page Table 5 3. Reference citations in text are corrected by section, with the affected beginning portions of the paragraphs republished under the appropriate section heading, followed by a listing of references to be added to the original published paper. (The following abstract of the original article appeared in record 2008-18378-010). We used a cross-sectional sample to compare ontogenetic trajectories in the concentrations of monoamine neurotransmitter metabolites in cerebrospinal fluid of wild anubis (Papio anubis, n = 49) and hamadryas (P. hamadryas, n = 54) baboons to test the prediction that they would differ, especially in males, in association with their distinct behavioral ontogenies. Values of all 3 metabolites [3-methoxy-4- hydroxyphenylglycol (MHPG), the norepinephrine metabolite; 5-hydroxyindoleacetic acid (5-HIAA), the serotonin metabolite; and homovanillic acid (HVA), the dopamine metabolite] declined consistently with dentally-calibrated maturation, and few taxon-related differences were apparent among juveniles. Adult females were too few for adequate comparison, but a discriminant function suggested that they might differ by taxon. Adult males of the 2 species differed strikingly from juveniles and from each other. Contrary to our initial hypothesis, adult male anubis had significantly lower HVA and MHPG, and higher 5-HIAA levels, than predicted from the overall, age-related trend, and MHPG continued to decline with age among adults. As young adults, male hamadryas had low 5-HIAA and a high HVA/5-HIAA ratio, while older males [normatively one-male unit (OMU) leaders] showed a reversal in the trend, with 5-HIAA rising and the HVA/5-HIAA ratio tending to fall. We speculate that the results are related to the dispersing and philopatric ontogenies of anubis and hamadryas males, respectively. Adult male anubis, whose fitness depends on building social networks with nonkin, have high relative serotonin activity, commonly associated with greater social circumspection and skill. Young adult male hamadryas, living among agnatic kin and mating opportunistically, exhibit low 5-HIAA levels, generally associated with impulsivity and social irresponsibility. This reverses as a male approaches the age at which he is normatively the leader of a one-male unit (OMU), and his fitness depends on his maintaining stable relationships with other leaders and with females.

The assembly line: Hexabenzocoronene amphiphiles appended with pyridyl-terminated triethylene glycol side chains, in combination with trans-[Pt(PhCN)(2)Cl(2)], lead to the formation of graphitic nanotubes. The structural features and dimensions of the nanotubes depend on the assembly conditions. A platinum(II)-bridged cyclic dimer having two HBC units self-assembles into a nanotubular structure.