The pallid sturgeon (Scaphirhynchus albus) is a federally endangered species, endemic to the Mississippi River drainage, stretching from the upper Missouri River in Montana to the Mississippi River, and continuing to the Gulf of Mexico. They are largely sympatric throughout this range with a close congener, the shovelnose sturgeon (S. platorhynchus), although speciation may have occurred when they were isolated in different refugia. In this study, we examined gene expression differences among pallid and shovelnose sturgeon families in response to heat stress. Gene expression can be considered a phenotype, and therefore, variability in expression can have an adaptive role in species. Additionally, we compared our results to a previous expression study that utilized RNA-Seq. We developed viable primer pairs for five genes in order to conduct RT-qPCR assays. There were significant differences in heat stress response between pallid and shovelnose sturgeon, potentially indicative of different evolved stress response pathways. Our species results contrasted with results from the previous study, indicating that further research is needed to improve the robustness of the results. Additionally, we found that offspring of hatchery and wild pallid sturgeon demonstrate different responses to heat stress, and potentially general stress that can occur in a hatchery environment. Overall, this study lays the groundwork for future research that can incorporate a larger suite of families to improve the robustness necessary to make actionable management recommendations.

Fertility is the most important factor controlling mammalian reproduction. Bull fertility, ability of the sperm to fertilize and activate the egg, and support embryonic development is crucial for early development. Similarly, the hormonal environment of the embryo also plays a critical role in successful embryonic development. We know that molecular health of the sperm as well as progesterone enhancing the development of the embryo is important for early development and implantation. The gaps in the knowledge base are 1) how early mammalian embryo development is paternally affected is not fully clear and 2) how progesterone improves the survival of the transferred embryo in the uterus still remains elusive. The central hypothesis was that low expression of MHC1 and Magel2 would cause a dysfunction in early embryo development and that progesterone will increase the survivability of the embryo via its associations with TNF-alpha and PGF2alpha. The objectives of this study were to 1) determine the expression of mhc1 and magel2 in single in vivo and in vitro blastocysts derived from low fertility and high fertility sires as well as to determine main pathways by which protein products of these genes regulate early development, and 2) identify the role of progesterone in regulating TNF-alpha and PGF2alpha. To accomplish these goals, we performed real time reverse transcriptase reaction and bioinformatics approaches, and ELISA and commercially available radioimmunoassay kits, respectively. The results of the experiments showed that 1) expression of mhcI transcripts were greater in high fertility in vivo embryos compared to their low fertility in vivo counterparts. Magel2 results showed an increase (P ≤ 0.05) in expression levels of high fertility in vivo embryos compared to their high fertility in vitro counterparts. Low fertility in vivo embryos had higher expression than high fertility in vitro embryos as well.

No description available.

Larval dispersal is one of the most complex phenomena that connects populations when larvae migrate over long distances from their native reefs to a new one. Dispersal outcomes are largely variable among individuals, even siblings, and the factors that drive this variability are not understood, making it challenging to create protection plans for marine species. Clownfish, Amphiprioninae, are among the most important coral reef species economically. Thus, creating protection plans for clownfish is necessary, especially as they are under habitat loss threat. Clownfish are considered an optimum model of organisms for studying larval dispersal and population connectivity mainly due to their relatively short larval dispersive phase and ease of breeding them in captivity. Since dispersal traits are influenced by genes, transcriptomic studies can reveal insights into larval dispersal. Thus, the variation in dispersal outcomes could be better understood if the differential expression of genes related to dispersal, such as swimming abilities, are considered. However, there is a lack of knowledge in the basics of gene expression in clownfish larvae. The aim in this thesis is to explore the ontogenic variability in RNA quantities across development in clownfish larvae as a first step of understanding the nature of their gene expression and optimizing RNA extractions and quantifications methods using clownfish as a model organism. We performed an RNA quantification study using larvae of Amphiprion bicinctus clownfish reared at KAUST SeaLabs. This thesis provides some insights regarding the pooled and individual quantities of RNA across time. The optimal pool size and age for various transcriptomic studies was determined. It was concluded that extracting RNA from young larvae is difficult. Depending on the research, it might be necessary to use older larvae or other techniques. The results from this thesis will build the foundation needed as a first step to perform differential gene expression studying the causes of variability in dispersal traits in A. bicinctus and the link between these traits and transcriptomics.

No description available.

No description available.

Ovarian secretions and growth hormone (GH) are major endocrine regulators of mammary growth and development in bovine mammary gland; but information on endocrine regulation during early prepubertal period is limited. Our overall objective was to study the regulation of mammary growth and development as well as transcript abundance in early prepubertal bovine mammary gland by ovarian secretions and exogenous bovine somatotropin (bST). In the first study, we determined the effect of staged ovariectomy on mammary growth and development in two to four month old Holstein heifers. In the second study, effects of bST on mammary growth and development, and also on putative stem cell population were evaluated by beginning bST treatment in one month old Holstein heifers. Mammary growth and development was determined by mass of mammary tissue, biochemical analyses, histological examination, transcript abundance and protein expression in mammary parenchyma (PAR) and fat pad (MFP). Ovariectomy reduced mass and lipid content of PAR without affecting the histological characteristics or rate of epithelial cell proliferation. There was a marked reduction in progesterone receptor expression both at the mRNA and protein level. Ovariectomy also reduced transcript abundance in GH receptor (R), insulin-like growth factor (IGF)-1, IGF binding protein -6, estrogen responsive and proliferation marker genes, but increased the mRNA abundance of IGF-1R in PAR. On the other hand, administration of bST did not have an impact on PAR growth, epithelial proliferation, putative stem cell population or transcript abundance of IGF-axis genes. However, bST reduced the relative abundance of GHR, signal transducers and activators of transcription-5b and suppressors of cytokine signaling-2 in mammary PAR. Transcript abundance of IGF-axis molecules, estrogen responsive genes and proliferation markers in MFP was not affected by ovariectomy or bST. Overall, our data suggest that ovary is a predominant regulator of mammary growth and development in prepubertal heifers and that exogenous bST is not effective as a mammary specific mitogen in very young prepubertal heifers. / Ph. D.

Classification of tissues and diseases based on gene expression data is a powerful application of DNA microarrays. Many popular classifiers like support vector machines, nearest-neighbour methods, and boosting have been applied successfully to this problem. However, it is difficult to determine from these classifiers which genes are responsible for the distinctions between the diseases. We propose a novel framework for classification of gene expression data based on notion of condition-specific clusters of co-expressed genes called xMotifs. Our xMotif-based classifier is biologically interpretable: we show how we can detect relationships between xMotifs and gene functional annotations. Our classifier achieves high-accuracy on leave-one-out cross-validation on both two-class and multi-class data. Our technique has the potential to be the method of choice for researchers interested in disease and tissue classification. / Master of Science

Diss. (sammanfattning) Stockholm : Karol. inst., 2006. / Härtill 4 uppsatser.

The transmission of Trypanosoma brucei, the causative agent of human African trypanosomiasis, depends upon the development in the bloodstream of 'stumpy forms' from non-transmissible 'slender forms'. In stumpy forms many mRNAs are downregulated and translation is generally repressed. However, a small subset of genes escape this repression and are upregulated, presumably as an adaptation for transmission. To understand the basic of this, regulatory sequences within the 3'UTR of a major stumpy-enriched transcript (an ESAG9 gene) have been characterised. This identified a signal responsible for gene silencing in slender forms and gene activation when cells develop to stumpy forms. An investigation was made of upstream open reading frames (uORFs) as a mechanism for the control of stumpy form gene expression. No evidence was found of uORF control, but one gene investigated was found to produce two transcripts through trans-splicing at different sites. These transcripts, which were found to exhibit some differential abundance between life-cycle stages, would generate a long and short form (from an internal ATG) of the encoded protein. Both are predicted to contain a UBA/TS-N (ubiquitin associated) domain, however, the longer form of the protein is also predicted to contain a transmembrane helix and cleavable signal peptide, suggesting a different localisation. However, ectopic expression of either protein form with a Ty epitope tag resulted in the same protein localisation. Additionally, the transcripts of two translational protein homologues, TbeIF4E4 and TbeIF6, were identified as upregulated in stumpy forms. Radiolabelled-methionine experiments and polysome analysis showed that overexpression or RNAi-mediated ablation of TbeIF6 resulted in a decrease in protein synthesis and decrease in translation. Unlike its archaeal homologue, TbeIF6 protein was not induced by coldshock treatment. Finally, to identify which transcripts escape translational repression in stumpy forms an analysis was made of polysome-associated transcripts by RNA-sequencing. This identified potentially interesting genes for further investigation, and showed that many procyclic-enriched transcripts were also enriched in stumpy form polysomeassociated RNA, confirming these cells as preadapted for transmission. Together, this work has characterised a 3’UTR regulatory element in a stumpy-enriched transcript, examined alternative trans-splicing of another transcript, investigated two translational protein homologues and identified transcripts that escape translational repression in the transmissible life-cycle stage of T. brucei.