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Effects of Dopamine Beta Hydroxylase Levels in a Mouse Model of FD
Get PDFFamilial dysautonomia is a severe, recessive disease that devastates the peripheral nervous system, culminating in death of most patients by age 40. The most debilitating feature of familial dysautonomia is the severe autonomic crises that occur. These crises, which can sometimes last for days, cause extreme vomiting and nausea, among other symptoms. The crises have been shown to coincide with an increased level of circulating dopamine following stress. The current hypothesis suggests that elevated levels of tyrosine hydroxylase cause an overproduction of dopamine. The chromaffin cells cannot convert this dopamine into norepinephrine quickly enough; therefore, this dopamine is released into the blood stream. We propose an alternate hypothesis in which the levels of dopamine beta hydroxylase are instead reduced. Reduction of dopamine beta hydroxylase, the enzyme that converts dopamine to norepinephrine, would result in a larger amount of dopamine being released from chromaffin cells during the response to stress. This reduction in enzyme levels is also seen in dopamine beta hydroxylase deficiency, a disease that shares many of the same symptoms of familial dysautonomia. In support of this hypothesis, we have shown through quantitative RT-PCR that dopamine beta hydroxylase transcript levels are decreased in Wnt1-Cre; IkbkapLoxP/ LoxP conditional knockout (CKO) embryos in which Ikbkap is ablated in the adrenal gland. Further analysis of the CKO using immunohistochemistry indicates that DBH protein levels may also be diminished as well as mis-localized within the cell
Coxiella Burnetii's Infection-Specific Small RNA 12 (CBSR12) Targets Cara and Metk Transcripts
Get PDFCoxiella burnetii is an obligate intracellular bacterial pathogen and the etiological agent of Q fever. Previous transcriptome analysis of C. burnetii by our group revealed several novel small RNAs (sRNAs) of varying sizes and expression patterns. Sequence alignments of these sRNA’s across all strains of C. burnetii show strong conservation, indicating a functional role for these RNA’s in C. burnetii’s intracellular lifestyle. Furthermore, during C. burnetii’s biphasic life cycle of metabolically active (LCV) and inactive (SCV) states, several of the sRNA’s have shown differential expression in SCV and LCV cells via RNA-Seq and Northern blot analyses. One such sRNA, termed CbsR12, showed a marked upregulation in infected Vero host cells when compared to bacteria grown in axenic media. Additionally, RNA-Seq data and qRT-PCR analyses show a marked upregulation of CbsR12 in LCV cells compared to SCV cells. Here, we show that C. burnetii RNase III cleaves CbsR12 into two fragments, an observation supported by both in silico and 5’ RACE analyses. in silico sRNA target prediction programs were used to determine possible mRNA targets of Cbsr12. We subsequently determined through in vitro electrophoretic mobility shift assays (EMSAs) and in vivo luciferase reporter assays, that CbsR12 binds carA transcripts, which codes for carbamoyl-phosphate synthase subunit A, and metK transcripts, which codes for S-adenosyl methionine synthetase. These genes code for essential enzymes involved in pyrimidine biosynthesis and the methionine cycle, respectively
Social Justice in the Library is Critical
Get PDFLibrarians have always been motivated by the needs of their patrons, and in today’s difficult political climate, we must address issues of social justice and equity. We have adjusted our practices to meet these needs, and will describe ideas for displays, programming, and curricula that have been successful in our libraries with the hope that you might be inspired toward social justice in yours. This paper will outline several areas where librarians can address social justice in connection to their communities
Elongator Function in the Anterior Pituitary and Its Relevance to Familial Dysautonomia
Get PDFFamilial Dysautonomia (FD) is a devastating neurodegenerative childhood disease characterized by a diminished number of autonomic neurons. FD children suffer from a multitude of autonomic symptoms including cardiovascular instability, gastrointestinal incoordination, and respiratory dysfunction. FD patients also exhibit an abnormal autonomic stress response, show poor growth velocity, and have difficulty gaining and maintaining weight. Treatment with growth hormone (GH) has been shown to increase growth velocity in FD patients. FD results from a mutation in the IKBKAP gene and diminished levels of the corresponding protein IKAP, a scaffolding protein that assembles a multi-subunit complex called Elongator. Elongator functions in the modification of tRNAs that mediate translation of AA- and AG-ending codons including lysine, glutamine, and glutamic acid. In the absence of Elongator, small AG biased genes are upregulated and large AA-biased genes are downregulated. IKAP is expressed throughout the autonomic nervous system and historically FD has been considered a strictly neurological disease. Here we show that IKAP is robustly expressed in the pituitary gland, indicating a strong dependence on Elongator. We hypothesize that compromised growth in FD may actually result from dysfunction of somatotrophs in the anterior pituitary, a non-neuronal cell type. To test this hypothesis, we generated a conditional knockout (CKO) mouse where Ikbkap is selectively ablated in anterior pituitary somatotrophs. These CKO mice exhibit decreased growth compared to control littermates. Surprisingly, quantitative immunohistochemistry indicates that GH1 levels may actually be increased in the CKO pituitary. CaBP7, a calcium binding protein that negatively regulates vesicle trafficking, is also found at elevated levels in the CKO, likely because of its strong AG-bias. In combination, these results suggest that upregulation of CaBP7 may inhibit GH1 exocytosis from pituitary cells, decreasing the amount of circulating GH1 and compromising growth in FD patients
Comparison & Analysis of Local Environmental Metagenomics and Diversity Sequencing Data Sets
Get PDFMetagenomics is the rapidly advancing field that studies genetic material extracted directly from environmental samples. Recent advances in computational and sequencing methodologies now give an array of options to biologists who seek to analyze their samples; however, these new technologies are often run independently with little comparison of the results obtained between methods on the same sample. Prior to this study, field work by local biology instructors in conjunction with the labs of Drs. Marisa Pedulla and Alysia Cox made use of two such techniques to analyze the DNA of a locally-obtained soil sample. One technique, “shotgun,” or metagenomic sequencing, sequenced all of the DNA molecules in the sample; the second technique, “diversity” sequencing, only sequenced the molecules amplified from a single gene, the 16S ribosomal subunit rRNA gene, of the prokaryotic DNA in the sample. Because the 16S gene is highly recognizable and species-specific, the latter method also provided a count of the occurrences of each prokaryotic species. Due to this feature, the use of the 16S diversity sequencing approach is commonly utilized for studies aiming to understand prokaryotic species representation in samples. When metagenomics DNA sequences are known, represented species and their number of occurrences in a sample may also be derived computationally by comparison to procured databases of known sequences of organisms. This exploratory study compared the commercially obtained results of bacteria proportions in our sample by the diversity method, along with two experimental computational methods using these sequences compared to the public databases. Our hypothesis was that metagenomics data would provide the most accurate portrayal of bacteria in the sample at the phylum taxon. Results found significant disparities in results between each method, with implications in microbiome studies of the environment and human gut
Determining the Place of AQP-3B in the WNT/CA2+ Noncanonical Pathway
Get PDFDuring Xenopus laevis gastrulation, convergent extension is required for the mesoderm to extend into the embryo and shape the embryonic body plan. Recent results from our lab suggest that the inhibition of aquaporin3b (aqp3b) prevents convergent extension of the mesoderm and that aqp3b acts through noncanonical Wnt signaling. Wnt signaling is a key signal pathway for embryo and tissue development and is composed of the canonical and the noncanonical pathways. Our lab has shown that aqp3b acts through a specific noncanonical pathway, the Wnt/Ca2+ pathway, and acts upstream of the cytoplasmic Wnt signaling pathway member Disheveled (Dsh). Frizzled7 (Fz7) is a membrane receptor in the noncanonical Wnt/Ca2+ pathway which also acts upstream of Disheveled. Our question for this project is whether aqp3b acts upstream or downstream of Fz7? When Fz7 was present, protein kinase C fused to green fluorescent protein (PKC-GFP) attached to the cell membrane but when Fz7 was absent, PKC-GFP remained freely in the cytoplasm. This served as our control injections: PKC-GFP + fz7 or PKC-GFP alone, respectively. The experimental injections included a morpholino (MO), small oligonucleotide that inhibited aqp3b expression, or a control MO which did not inhibit aqp3b. We have shown that the MO has kept PKC-GFP localized to the cytoplasm despite the presence of Fz7, while the control MO is allowing PKC-GFP membrane attachment. These data demonstrate that aqp3b acts downstream of the Fz7 receptor in the noncanonical Wnt signal cascade
Assessment of the Occurrence and Distribution of Unpermitted Wastewater Treatment Systems Along a Section of Bozeman Creek and Identification of Recommendations to Address Fecal Contamination
Get PDFIntroduction - Bozeman Creek is considered “impaired” because sediment, nutrients and E. coli are at levels that impair the use of water for beneficial purposes such as irrigation or recreation. These E. coli can come from wildlife, livestock, pets or leaking septic tanks. Previous monitoring points to wastewater from septic systems as a major source of contamination to this stream. The research goal is to assess the impact of unpermitted wastewater systems on Bozeman Creek and to identify recommendations to address aging septic systems.
Methods - Online Health Department records are being reviewed to research the following questions: 1) What percentage of septic systems along a defined area of Bozeman Creek are unpermitted and what are their locations? 2) What percentage of permitted systems along a defined area of Bozeman Creek are >25 years old, and what are their locations? Additionally, a literature search is being conducted to identify best practices for the department and partners to address the status of onsite wastewater treatment along Bozeman Creek. GIS model is being created to assess which properties are most at risk of contaminating the creek.
Results - 34 properties were identified along the portion of Bozeman creek; 47 % of these properties lack septic system permits. Of those with permits, 56% are systems 25 years or older. Development of a GIS risk assessment model is underway.
Conclusions - The results of this project will be useful to health department staff to address wastewater sources contaminating Bozeman creek and serve as a model for other waterways
Maternal Effects of Aseptic and Septic Injury on Embryonic Larval Gene Expression in the Tobacco Hornworm, Manduca Sexta
Get PDFCross-generational effects of physical and pathogenic stress have been demonstrated in several insect groups, including our model insect Manduca sexta. Prior studies in our laboratory have shown that maternal exposure to the soil-dwelling gram-negative bacteria, Serratia marcescens, just prior to adult eclosion alters egg morphology and larval immunity. Our goal is to identify mechanisms underlaying pathogen-associated parental effects on offspring. The current study advances this goal through measurement of embryonic size, embryonic histone modification, and both embryonic and larval gene expression. Two days prior to eclosion, parents were injected with saline, heat killed S. marcescens, or live S. marcescens. Embryos were collected at 24 (+/- 2) h or permitted to hatch for clearance assays (first instar) or measurement of fat body gene expression (fourth instar). We find that maternal, but not paternal, pathogen exposure significantly increases egg volume variability, and that maternal pathogen exposure may delay hatching. Furthermore, maternal injection with bacteria conferred on their offspring an enhanced ability to clear infection when compared to their saline injected peers. Histone analysis revealed that maternal treatment does not globally alter embryonic histones, however, several immune-related genes demonstrated altered expression in both embryos and fourth instar larvae
In the Interim: Leadership Shorts from Three Interim Library Directors
No full textLeadership can take many forms. For some it is leading by example, while for some it is more authori-tarian, and yet others may have a team leadership model. Whatever the leadership model is, how does it change in a short term appointment? This article gives three perspectives from interim library leaders at different institutions. One is a large state university, one a private college, and one a smaller state uni-versity with a tech focus. While the institutions and the circumstances of the interim roles may be dif-ferent, there are similarities that cross all three