Browsing by Subject "gene expression"
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(2019)Colorectal cancer (CRC) kills more than half a million people a year worldwide. Usually the disease develops over several years via multiple steps which involve both genetic and epigenetic alterations. CRC is often diagnosed at late stage, when the cancer has already metastasized, and the prognosis is relatively poor. Several studies suggest that the first changes towards colorectal cancer occur and can be detected in histologically normal tissue before the appearance of any detectable lesion. The precancerous cells harbouring those changes may form a field of tissue, which is predisposed to malignant transformation. The study of pre-cancerous tissue might reveal the earliest changes in CRC development, which can be used as biomarkers for early detection and prevention of CRC. The aim of this thesis was to revise and investigate whether the aberrant expression of the six chromosomal segregation genes, Bub1, Mis18a, Pms2, Rad9a, Tpx2, and Mlh1, would signal carcinogenesis in mouse colon mucosa. Altogether fourteen mice, of which six had a proximal colon carcinoma, were selected for the study. The expression analysis was performed to histologically normal colon mucosa collected from the proximal and distal colon of each mice in order to investigate whether the possible pre-cancerous changes are found exclusively in the close proximity to the carcinoma. The expression was quantified with reverse transcription quantitative polymerase chain reaction (RTqPCR). No statistically significant gene expression differences were found between the carcinoma and control mice, indicating that the studied mice did not display cancer-preceding expression changes of the six studied genes in the carcinoma adjacent histologically normal colon mucosa. The results differed from the previously reported results, where the expressions of the six genes were found to be downregulated in the carcinoma adjacent mucosa. Here, the sample size was presumably not large enough to reveal statistically significant clustering of the expression patterns. However, Bub1 seemed to have a downregulated trend in the carcinoma adjacent mucosa, which supports the previously suggested role of Bub1 alterations in CRC initiation.
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(2019)Human umbilical vein endothelial cells are responsible for maintaining and forming new vessels from existing ones, in a biological process called sprouting angiogenesis. Sprouting angiogenesis is a crucial mechanism for the resolution of hypoxia and normal development of tissues. It also plays a key role in internal plague hemorrhages, which can lead to embolisms and other cardiovascular complications. Angiogenesis is also crucial for cancer development. Sprouting angiogenesis is initiated by hypoxic tissue excreted vascular endothelial growth factor gradient, which induces normal endothelial cells into either a proliferative stalk cell or a signal sensing tip cell phenotype. Both of these cell types depend on the rapid flow of lipids to their plasma membrane, either to form plasma membrane protrusions in tip cells or as new plasma membrane material in dividing stalk cells. This flow is envisioned to involve both vesicle-mediated and non-vesicular mechanisms. A major non-vesicular route of lipid transfer occurs at membrane contact sites via lipid transport proteins. Furthermore, lipids can be transported to the plasma membrane by the direct fusion of vesicles or endosomes with the plasma membrane This thesis set out to explore the role of two membrane contact site proteins, oxysterol-binding protein- related protein 2 and protrudin, in angiogenesis and lipid transfer. Their role was examined by RNA-sequencing transient knock-down samples of these proteins in HUVECs. The RNA-sequencing data was examined by differential expression, gene ontology overrepresentation and gene set enrichment analyses. Gene expression analysis provided almost 10 000 significantly changed transcripts (adjusted p-values < 0.05), in each silenced cell type. The distribution of differentially expressed genes in oxysterol-binding protein- related protein 2 silenced cells, is skewed toward negative fold changes, whereas the distribution of differentially expressed genes in protrudin silenced samples is normally distributed. The results also show significant changes in gene ontologies related to proliferation, cell cycle, angiogenesis as well as hypoxia in both sample types. Gene set enrichment analysis showed upregulation in angiogenesis related pathways, such as the PI3K-Akt and MAPK pathways, in both samples. Significant downregulation was present in cell cycle related pathways and cholesterol biosynthesis pathway in both ORP2 and protrudin silenced samples.
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(2020)Social insects such as ants live in societies and have a strict division of labor between reproductive and worker castes. A colony can consist of even millions of individuals and the number of queens can vary a lot. Populations where each colony comprises just one or few queens are often called kin structured because the relatedness between nestmates is high. Colonies that have lots of queens and the society lives in many connected nests (polydomy) in are referred to as supercolonies. In these colonies relatedness between individuals is low and the workers represent many genetic lineages. Depending on species and the environment where the colony lives societies can behave aggressively towards individuals from other nests to protect their own nest. Ants must be able to recognize members of their own colony from the intruders to be able to protect the nest. Nestmate recognition is a key element in the interaction between nests and species and makes it possible for the workers in the colony to favour their own nestmates in form of care, defence or food acquisition to gain inclusive fitness benefits. To recognise nestmates ants must be able to sense chemical cues. Ants detect these chemical signals through the proteins expressed mainly in their antennas. In this thesis I studied gene expression of genes related to chemosensation in seven Formica species using the RT qPCR method. My study species were kin structured Formica exsecta, F. pratensis and F. fusca and supercolonial F. truncorum, F. pressilabris, F. cinerea and F. aquilonia. My study genes belong to gene families that code for odorant binding proteins (OBP), chemosensory proteins (CSP) and gustatory reseptors (GRT). I want to find out whether the expression of these genes differs between castes, and whether the caste difference varies between kin structured and supercolonial species. Workers have many tasks in the ant colony and to take care of them, they need to have a sophisticated sensory system. For that reason, I expect to find out that the study genes are expressed more in the worker than the queen caste. In addition, I expect the caste difference in gene expression to be higher in the kin structured species than in the supercolonial species. That is because kin structured species behave more aggressively towards intruders and possibly confront intruders more often than the individuals living in supercolonies. Furthermore, in the supercolonies low relatedness between individuals sometimes lead to conflicts inside the nest. For that reason, I suppose queens of the supercolonies express chemosensory genes more than the queens from the kin structured colonies. Overall expression level was the highest for the OBP and the lowest for GRT. The expression level of CSP was in between these extremes. In accordance with my hypothesis gene expression of OBP and CSP was higher in workers in all the study species. GRT expression was worker biased in six of the seven species. Caste difference in expression of chemosensory genes was similar in kin structured and supercolonial species. The expression level varied between species but did not show a pattern depending on the degree of the polygyny. The study revealed that the expression of OBP and CSP is correlated. My results revealed expected worker biased pattern in the expression. The result might be a consequence of better olfactory or taste abilities in the worker caste compared to queens or it may even be consequence of more sophisticated nestmate recognition skills of the workers. This study reveals valuable information about the gene expression of chemosensory genes related to the recognition system in the ants and awakes many new study questions. Chemical sensory system has been studied a lot in the ants, but in the field of expression studies there is still lot to reveal.
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(2022)Gravity has a modifying effect on plant architecture. The phytohormone auxin is known to transmit the signal of gravity perception from gravity-sensing cells to responsive tissues and cause an asymmetric growth response in the receiving organs. Intercellular auxin flux is mediated by many different transporter proteins, of which PIN-FORMED 3 (PIN3) is known to function as an auxin efflux carrier in gravitropic responses. The expression of PIN3 is known to locate in one cell layer of the shoot endodermis in herbaceous plant species and Populus hybrids. The objective of this study was to determine the location of PIN3 ortholog expression using silver birch (Betula pendula) as a model plant of a woody plant species. Agrobacterium tumefaciens C58 strain GV3101 (pMP90), harbouring binary vector pBpPIN3-erVEN-nosT2 containing erVenus (erVEN-YFP) as a marker gene under the BpPIN3 promoter and terminator nosT2, was used in the study to detect the gene expression. The expression vector was constructed by the Gateway® cloning method and transformed into in vitro shoot explants of silver birch by Agrobacterium-mediated transformation (AMT). In the study, transgenic lines were generated successfully, from which the fluorescence of erVEN was observed in the cross-section of the shoot with a fluorescence stereo microscope. The transgenicity of putative transgenic lines was confirmed by PCR of erVEN. BpPIN3 was likely to be expressed to endodermal tissue in the shoots of silver birch. This study can be considered as a screening step for the localization of BpPIN3 expression. The study facilitates the discovery of factors related to the regulation of PIN3 in tropic responses in woody plant species. This information can in the future be utilized in plant breeding to optimize plant architecture.
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