Master of Science - Adult Education / Mutations were sought in Escherichia coli ribosomal RNA and ribosomal proteins that confer dependence to the antibiotic streptomycin, using both newly available as well as well-established genetic systems. I found that a classical ribosomal mutant, Sm-D3, was streptomycin dependent and had an additional mutation in another ribosomal component – protein L7/L12. The double mutant had an 8-fold lower streptomycin requirement as compared to Sm-D3 with a wild-type rplL. This supported a functional involvement of L7/L12 in the decoding center of the ribosome.

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

Familial lipoprotein lipase (LPL) deficiency is a rare disorder of lipid metabolism associated with massive chylomicronaemia. Patients often present early in life with abdominal pain, pancreatitis, hepatosplenomegaly, eruptive xanthomata and zero to near zero levels of LPL activity in post-heparin plasma. The genetic heterogeneity underlying this disease is well-characterised and over 40 mutations have been described at the LPL gene loci. In this report three mutations are described at the LPL locus in two unrelated probands, namely, JJ (Kindred I) and LB (Kindred II). JJ presented early in childhood with signs and symptoms suggestive of LPL-deficiency. These were abdominal pain, hepatosplenomegaly and a markedly reduced LPL activity (38% of normal) in post-heparin plasma. DNA studies showed JJ to be a compound heterozygote for two point mutations in the LPL gene, these being, the I194T and C418Y substitutions, which occur in exons 5 and 9, respectively. Several mutation detection systems were set up as part of the characterisation and screening workup for these mutations; these were, allele-specific oligo nucleotide (ASO) hybridisation, "ARMS" PCR, PCR-SSCP, RT-PCR and DNA sequence analysis. In an earlier separate study, in vitro transfection results showed that the I194T mutant was catalytically inactive. Our findings of zero LPL activity in JJ's post-heparin plasma, implies that the C418Y mutation is also likely to produce an inactive protein product. The differences in LPL activity observed during the pre- and post-pubertal stages, if not artefactual, may be due to differential processing of LPL during human development with loss of activity post puberty. LB was first diagnosed with pancreatitis during the third trimester of her pregnancy. Although her child, BB, was successfully delivered by caesarean section, LB died of haemorrhagic pancreatitis with the marked hyperlipidaemia being suggestive of an underlying deficiency in LPL activity. Genomic DNA from her parents was first subjected to mutation analysis, since only slide specimens of post-mortem material were available from LB. Maternal DNA revealed a 0-A transition at nucleotide position 1516 which results in the substitution of lysine for glutamic acid at codon 421 in exon 9 (E421K), while paternal DNA show a single polymorphism at codon 108 in exon 3 of the LPL gene. Analysis of archival DNA obtained from histopathological slides of spleen tissue from LB also showed the E421K mutation. This mutation was also detected in her offspring, BB indicating maternal inheritance in three generations. While, this mutation may produce a catalytically defective product, the evidence is insufficient to propose a role for LPL deficiency as the primary cause of death in this patient, hence the search for a second mutation in the LPL gene of LB is imperative to establish this association.

Influenza is a seasonal viral disease affecting over 1 billion people annually around the globe, as reported by the World Health Organization (WHO). The influenza virus has been around for decades causing multiple pandemics and encouraging researchers to perform extensive analysis of its evolutionary patterns. Current research uses phylogenetic trees as the basis to guide population genetics and other phenotypic characteristics when describing the evolution of the influenza genome. Phylogenetic trees are one form of representing the evolutionary trends of sequenced genomes, but that do not capture the multidimensional complexity of mutational pathways. We suggest representing antigenic drifts within influenza A/H1N1 hemagglutinin (HA) protein as a graph, $G = (V, E)$, where $V$ is the set of vertices representing each possible sequence and $E$ is the set of edges representing single amino acid substitutions. Each transition is characterized by a Malthusian fitness model incorporating the genetic adaptation, vaccine similarity, and historical epidemiological response using mortality as the metric where available. Applying reinforcement learning with the vertices as states, edges as actions, and fitness as the reward, we learn the high likelihood mutational pathways and optimal policy, without exploring the entire space of the graph, $G$. Our average predicted versus actual sequence distance of $3.6 \pm 1.2$ amino acids indicates that our novel approach of using naive Q-learning can assist with influenza strain predictions, thus improving vaccine selection for future disease seasons. / Master of Science / Influenza is a seasonal virus affecting over 1 billion people annually around the globe, as reported by the World Health Organization (WHO). The effectiveness of influenza vaccines varies tremendously by the type (A, B, C or D) and season. Of note is the pandemic of 2009, where the influenza A H1N1 virus mutants were significantly different from the chosen vaccine composition. It is pertinent to understand and predict the underlying genetic and environmental behavior of influenza virus mutants to be able to determine the vaccine composition for future seasons, preventing another pandemic. Given the recent 2020 COVID-19 pandemic, which is also a virus that affects the upper respiratory system, novel approaches to predict viruses need to be investigated now more than ever. Thus, in this thesis, I develop a novel approach to predicting a portion of the influenza A H1N1 viruses using machine learning.

B-cell chronic lymphocytic leukemia is the most common leukemia in the adult population of Western developed countries. In 2005, an estimated 9,730 adults in the United States will be diagnosed with B-CLL and an estimated 4,600 deaths will occur. B-CLL is a common heterogeneous malignant disease with variable outcome. B-CLL is divided into two groups based on whether somatic hypermutation is observed in the variable region of the immunoglobulin heavy-chain locus (IgVH). The two distinct groups are named mutated and unmutated. The B-CLL mutated group has a more favorable prognosis than the unmutated group. Gene expression profiling has been used successfully to decipher the biological and clinical diversity of many leukemias and lymphomas. Recently, other small RNAs (microRNAs) have been shown to be important in hematopoiesis. MicroRNAs are small 20-28 nucleotide RNAs that are believed to control many important cellular and developmental processes by posttranscriptional gene silencing, translational repression, and modulating epigenetic events. We are interested in whether microRNA expression correlates with the mutational status of IgVH. This study is significant in the following ways: (1) microRNAs may become surrogate markers for the mutational status of IgVH of B-CLL, which implies a more rapid diagnostic means as compared to the current practice, and (2) microRNAs, in the particular context of B-CLL, may play some significant roles in a gene regulatory network that is further responsible for chromosomal abnormalities found in B-CLL. This thesis presents a study comparing microRNA expression in mutated and unmutated B-CLL groups. Instead of using a genome-wide expression profiling strategy, we selected a specific set of microRNAs based on their chromosome locations and mRNA targets. Specifically, we chose the following eight microRNAs (with their chromosomal abnormalities): mir16-1 (deletion 13), let-7i (trisomy 12), mir196-2 (trisomy 12), mir26a-2 (trisomy 12), mir-34b (deletion 11), mir-125b (deletion 11), mir-181C (trisomy 19), mir-125a (trisomy 19). We used solution hybridization assays to monitor the expression of microRNAs. We successfully characterized the microRNA expression in twelve B-CLL patient samples (eight mutated and four unmutated). Among the eight microRNAs examined, three (mir196-2, mir-125a, mir-125b) are not expressed in the two B-CLL groups, four (mir16-1, mir26a-2, let-7i, mir-34b) have significant differences in expressions over the two groups, and one (mir-181c) has no significant difference in expressions over the two groups.

B-cell chronic lymphocytic leukemia is the most common leukemia in the adult population of Western developed countries. In 2005, an estimated 9,730 adults in the United States will be diagnosed with B-CLL and an estimated 4,600 deaths will occur. B-CLL is a common heterogeneous malignant disease with variable outcome. B-CLL is divided into two groups based on whether somatic hypermutation is observed in the variable region of the immunoglobulin heavy-chain locus (IgVH). The two distinct groups are named mutated and unmutated. The B-CLL mutated group has a more favorable prognosis than the unmutated group. Gene expression profiling has been used successfully to decipher the biological and clinical diversity of many leukemias and lymphomas. Recently, other small RNAs (microRNAs) have been shown to be important in hematopoiesis. MicroRNAs are small 20-28 nucleotide RNAs that are believed to control many important cellular and developmental processes by posttranscriptional gene silencing, translational repression, and modulating epigenetic events. We are interested in whether microRNA expression correlates with the mutational status of IgVH. This study is significant in the following ways: (1) microRNAs may become surrogate markers for the mutational status of IgVH of B-CLL, which implies a more rapid diagnostic means as compared to the current practice, and (2) microRNAs, in the particular context of B-CLL, may play some significant roles in a gene regulatory network that is further responsible for chromosomal abnormalities found in B-CLL. This thesis presents a study comparing microRNA expression in mutated and unmutated B-CLL groups. Instead of using a genome-wide expression profiling strategy, we selected a specific set of microRNAs based on their chromosome locations and mRNA targets. Specifically, we chose the following eight microRNAs (with their chromosomal abnormalities): mir16-1 (deletion 13), let-7i (trisomy 12), mir196-2 (trisomy 12), mir26a-2 (trisomy 12), mir-34b (deletion 11), mir-125b (deletion 11), mir-181C (trisomy 19), mir-125a (trisomy 19). We used solution hybridization assays to monitor the expression of microRNAs. We successfully characterized the microRNA expression in twelve B-CLL patient samples (eight mutated and four unmutated). Among the eight microRNAs examined, three (mir196-2, mir-125a, mir-125b) are not expressed in the two B-CLL groups, four (mir16-1, mir26a-2, let-7i, mir-34b) have significant differences in expressions over the two groups, and one (mir-181c) has no significant difference in expressions over the two groups.

No description available.

All cancers were once normal cells. They became cancerous through the chance acquisition of particular somatic mutations that gave them a selective advantage over their neighbours. Thus, the mutations that initiate cancer occur in normal cells, and the normal clonal dynamics of the tissue determine a mutant cell's ability to establish a malignant clone; yet these remain poorly understood in humans. One tissue was selected for the exploration of each of these two facets of somatic evolution: blood for clonal dynamics; colon for mutational processes. Blood presents an opportunity to study normal human clonal dynamics, as clones mix spatially and longitudinal samples can be taken. We isolated 140 single haematopoietic stem and progenitor cells from a healthy 59 year-old and grew them in vitro into colonies that were whole genome sequenced. Population genetics approaches were applied to this dataset, allowing us to elucidate for the first time the number of active haematopoietic stem cells, the rate at which clones grow and shrink, and the cellular output of stem cell clones. Colonic epithelium is organised into crypts, at the base of which sit a small number of stem cells. All cells in a crypt ultimately share an ancestor in one stem cell that existed recently, and consequently share the mutations that were present in this ancestor. We exploited this natural clonal unit, isolating single colonic crypts through laser capture microdissection. 570 colonic crypts from 42 individuals were whole genome sequenced. We describe the burden and pattern of somatic mutations in these genomes and their variability across and within different people, identifying some mutational processes that are ubiquitous and others that are sporadic. Targeted sequencing of an additional 1,500 crypts allowed us to quantify the frequency of driver mutations in normal human colon. Together, these two studies inform on the somatic evolution of normal tissues, describing new biology in human tissue homeostasis and providing a window into the processes that govern cancer incidence.

Haemonchus contortus is a parasitic nematode that is controlled in large part by nematocidal drugs that target receptors of the parasitic nervous system. Hco-UNC-49 is a nematode GABA receptor that has a relatively low overall sequence homology to mammalian GABA receptors but is very similar to the UNC-49 receptor found in the free living nematode Caenorhabditis elegans. However, the nematode receptors do exhibit different sensitivities to GABA which may be linked to differences in the putative GABA binding domains. Mutational analysis conducted in this study identified at least one amino acid, positioned near the GABA binding domain, which may partially account for differences in nematode GABA sensitivity. In addition, positions reported to be crucial for GABA sensitivity in mammalian receptors also affect GABA sensitivity in Hco- UNC-49 suggesting that the GABA binding domains of the mammalian and nematode GABA receptors share some pharmacological similarities. However, there were some differences observed. For example, in mammalian GABAA receptors amino acids from both  and  subunits appear to be important for GABA sensitivity. For residues examined in this study, only those on the UNC-49B subunit, and not UNC-49C, appear important for GABA sensitivity. / UOIT

Diss. (sammanfattning) Linköping : Linköpings universitet, 2007. / Härtill 4 uppsatser.