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Publication GENETIC CONNECTIVITY OF FOUR MANGROVE SPECIES FROM THE MALAY PENINSULA(2013-08-12) WEE KIM SHAN; BIOLOGICAL SCIENCES; WEBB, EDWARD LAYMANAs habitat fragmentation and degradation continues to threaten the integrity of genetic connectivity in mangroves, gene flow among populations becomes increasingly important for the maintenance of sufficient genetic diversity and evolutionary potential. In this thesis, I determined the effects of reproductive traits, physical barriers and ocean currents on gene flow in mangroves, by examining the genetic connectivity of four major mangrove species from the Malay Peninsula. My data revealed that the relative genetic connectivity across species reflects their relative propagule dispersal potential, indicating that dispersal traits have a significant influence over gene flow. Comparative analysis across species demonstrated that the peninsula was a differential barrier to gene flow, and that ocean currents are important in defining genetic connectivity. Understanding gene flow in mangroves and the intrinsic and extrinsic factors behind it is crucial in assessing the genetic susceptibility of mangroves towards and the potential of recovery from habitat disturbances.Publication DEVELOPMENT AND CHARACTERIZATION OF DNA APTAMERS THAT TARGET GLUTEN PEPTIDES RESPONSIBLE FOR CELIAC DISEASE DEVELOPMENT(2013-08-22) ROOPSHA BRAHMA; BIOLOGICAL SCIENCES; KIM CHU-YOUNGCELIAC DISEASE IS TRIGGERED BY EXPOSURE TO WHEAT GLUTEN AND SIMILAR PROTEINS FOUND IN BARLEY AND RYE. WE SELECTED TWO SETS OF SINGLE STRAND DNA APTAMERS THAT TARGET THE IMMUNODOMINANT 33-MER GLUTEN EPITOPE AND INVESTIGATED THEIR ABILITY TO PREVENT GLUTEN-INDUCED AUTOIMMUNITY. THE FIRST SET OF APTAMERS, WHICH BIND TO THE NATIVE 33-MER GLUTEN PEPTIDE (LQLQPFPQPQLPYPQPQPLYPQPQLPYPQPQPF), INHIBITS TISSUE TRANSGLUTAMINASE-MEDIATED DEAMIDATION WHICH IS A NECESSARY STEP IN CELIAC DISEASE DEVELOPMENT. THE SECOND SET OF APTAMERS, WHICH BIND TO THE DEAMIDATED 33-MER GLUTEN PEPTIDE (LQLQPFPQPELPYPQPEPLYPQPELPYPQPQPF), HINDERS EPITOPE LOADING ONTO HLA-DQ2. THESE APTAMERS MAY SERVE AS A PREVENTIVE THERAPEUTIC FOR CELIAC DISEASE.Publication DIVERSE WAYS OF CONTROLLING miRNA EXPRESSION DURING FLY DEVELOPMENT(2017-01-16) ZHOU LI; BIOLOGICAL SCIENCES; OKAMURA KATSUTOMO; TUCKER-KELLOGG, GREGAs significant gene regulators, miRNAs are processed from long mRNA-like transcripts (primary miRNAs) by sequential actions of ribonucleases Drosha and Dicer. But how the expression of miRNAs could be controlled at each step is currently unknown. To explore the underlying mechanisms, we did expression profiling analyses on mature and primary miRNAs from small and long RNA-seq libraries, respectively, which were generated from a time course covering the whole Drosophila embryogenesis. Although majority miRNAs generated from the same polycistronic transcript, also called ‘clustered miRNAs’, tended to show similar expression patterns, dissimarility in expression changes between individual members of clusters was also observed, indicating that the transcription level was not the only determinant. Further analyses showed that distinctive miRNA stabilities, alternative promoter usage and differential Drosha cleavage efficiency were some of the bases of the differential regulation of miRNA clusters, demonstrating that miRNAs were regulated by diverse mechanisms.Publication Genetic control of virulence in Erwinia chrysanthemi pv. Zeae(2008-01-28) MUMTAZ BEGUM BINTE MOHAMED HUSSAIN; BIOLOGICAL SCIENCES; ZHANG LIANHUIThis study focuses on a bacterial isolate EC1 from rice showing foot rot symptoms. Genetic and phenotype analysis show that EC1 is a member of E. chrysanthemi but displays a unique ability to inhibit rice seeds germination, suggesting it belongs to a rarely characterized pathovar E. chrysanthemi pv. zeae. Transposon disruption of the expI gene encoding the biosynthesis of the quorum sensing signal OHHL significantly enhances the bacterial cell motility, decreases its virulence on potato, and marginally affects its inhibitory activity on rice seeds germination. Further study reveals that EC1 produces an antibiotic-like toxin, which inhibits rice seeds germination. Subsequent mutagenesis and DNA sequence analysis has identified several genes encoding toxin production and regulation. The toxin defective mutants lost the inhibitory activity on rice seeds. These data show for the first time that a novel toxin is a key virulence factor of E. chrysanthemi pv. zeae.Publication Structural and epitope characterization of dust mites allergens, Der f 13 and Blo t 5(2007-05-04) CHAN SIEW LEONG; BIOLOGICAL SCIENCES; MOK YU KEUNG, HENRYThe structure of Der f 13, a group 13 allergen from dust mite Dermatophagoides farinae was solved using NMR spectroscopy. Epitope residues have been identified and mapped to Glu-41, Lys-63, Lys-91 and Lys-103. A triple mutant 3A (E41A_K63A_K91A) has been generated and shown to stimulate the release of Th1 cytokines in patientsa?? PBMC while at the same time reducing the secretion of Th2 cytokines. The 3A mutant is able to stimulate mouse IgG antibodies that are able to inhibit the binding of patientsa?? IgE to wild type Der f 13, indicating its potential as candidate for vaccine immunotherapy against Der f 13 allergy. Structural characterization of Blo t 5, from Blomia tropicalis has also been carried out using CD and NMR spectroscopy. Backbone chemical shifts of Blo t 5 have also been assigned and three I?- helices separated by tight turns were observed in the chemical shift index plot.Publication Roles of BPGAP1 in cell signalling - Filling in the "GAP" in cell dynamics control(2005-10-03) LUA BEE LENG; BIOLOGICAL SCIENCES; LOW BOON CHUANRHO GTPASES ARE IMPORTANT REGULATORS FOR CELL DYNAMICS. THEY ARE ACTIVATED BY GUANINE NUCLEOTIDE EXCHANGE FACTORS AND INACTIVATED BY GTPASE-ACTIVATING PROTEINS (GAPS). WE RECENTLY IDENTIFIED BPGAP1 AS A NOVEL RHOGAP THAT REGULATES PSEUDOPODIA AND CELL MIGRATION VIA THE INTERPLAY OF ITS BNIP-2 AND CDC42GAP HOMOLOGY DOMAIN, RHOGAP DOMAIN AND PROLINE-RICH REGION. TO FURTHER ELUCIDATE ITS ROLES IN CELL DYNAMICS CONTROL, WE EMPLOYED PROTEIN PRECIPITATIONS AND MATRIX-ASSISTED LASER DESORPTION/IONIZATION MASS-SPECTROMETRY AND IDENTIFIED CORTACTIN AND EEN/ENDOPHILIN II AS NOVEL PARTNERS OF BPGAP1. PROGRESSIVE DELETION AND SITE-DIRECTED MUTAGENESIS STUDIES CONFIRMED THAT THEY INTERACTED DIRECTLY WITH THE PROLINES AT 184 AND 186 OF BPGAP1 VIA THEIR SRC HOMOLOGY-3 DOMAIN. WE FURTHER SHOWED THAT BPGAP1 INTERACTION WITH CORTACTIN FACILITATED CORTACTIN TRANSLOCATION TO THE PERIPHERY FOR ENHANCED CELL MIGRATION AND THAT THE OVEREXPRESSION OF EEN AND BPGAP1 ENHANCED EGF-STIMULATED RECEPTOR ENDOCYTO
Publication VARIATION IN PRIMATE EYE COLORATION AND MORPHOLOGY: PHYLOGENETIC, SOCIO-ENVIRONMENTAL, AND FUNCTIONAL PERSPECTIVES(2021-08-16) JUAN OLVIDO PEREA GARCIA; BIOLOGICAL SCIENCES; Fraser Monteiro, Antonia AlbertoExternal eye appearance is vastly diverse and colorful in primates, but we do not know what this splendid variation responds to. Early studies suggested that human eyes evolved to stand out, while all other primates evolved dull eyes to go unnoticed, leading to the proposal that human eyes evolved to communicate in human-unique ways. Our current understanding of external eye appearance is at odds with this “conspicuous-cryptic” dichotomy - recent studies suggest that external eye appearance in some nonhuman primates may be as salient as our own. If communicative functions don’t drive diversity in external eye appearance, what does? Here, I quantify external eye variation in different primate taxa to test hypotheses about its functions and origins. Most results suggest that light is an important driver of external eye appearance in primates that has received surprisingly little attention, given that eyes mediate an organisms’ relationship with its environment through light.Publication Expression and characterization of FAT1 and atrophin 1 proteins regulating planar cell polarity and MBD1 protein involved in lymphoma(2009-02-04) ANUPAMA VAASUDEVAN; BIOLOGICAL SCIENCES; KUNCHITHAPADAM SWAMINATHANFat, the first tumor suppressor gene to be discovered in Drosophila melanogester, is one of the most important regulators of planar cell polarity which controls the directional alignment of hair bristles and photoreceptors in the eyes of Drosophila. The mammalian counterpart of Fat known as Fat1 has been found to play a vital role during cerebral development, glomerular slit formation and gastrulation. Atrophin1 (also known as grunge) is a nuclear receptor which is predominately found in the nucleus but sometimes shuttles to the cytoplasm. The C-terminus of Atrophin is shown to interact with the C-terminal domain of Fat in the regulation of planar cell polarity. The precise role of these two important molecules in planar cell polarity is yet to be fully understood. Apart from its role in the Fat-Atrophin complex, Atrophin1 like proteins have been implicated in Dentatorbral Pallidoluysian Atrophy, a neural disorder. The structures of the C-terminal domains of Fat1 (160 aa) and Atrophin1 (196 aa) from Mus musculus (to be solved, separately and for their complex, using X-ray crystallography) will provide a pedestal for understanding the roles of Fat1 and Atrophin1 in the mechanism of regulation in planar cell polarity. MBD1 or Methyl binding domain 1 protein belongs to the class of Methyl CpG binding proteins (MBD 1-4 and MeCP2).The sequence similarity of these proteins is restricted only in their MBD domain, thus highlighting different roles. MBD1 has additional TRD and Zinc finger domains, which bind to non-methylated DNA and silence them, while the MBD domain silences hypermethylated DNA. The dual DNA binding capacity of MBD1 is of great importance in understanding tumorigenesis, very little of which is currently known. The solution structure of the human MBD domain in complex with DNA has been solved. Currently, we are cloning full length MBD1 (605 aa) from a human lymphoma cell line into the p Fast Bac Htb vector for baculovirus expression, after failure to express this protein in bacterial expression systems using the pET14b and pET32A vectors.Publication Study different scl isoforms in zebrafish primitive and definite hematopoiesis(2008-11-24) ZHEN FENG HUA; BIOLOGICAL SCIENCES; WEN ZILONGHematopoiesis is a highly organized program composed of ventral mesoderm induction, hematopoietic stem cell (HSC) specification and subsequent lineage differentiation. The SCL gene, encoding a basic helix-loop-helix (bHLH) transcription factor, has been indicated important for the development of primitive and definitive hematopoiesis. In this research, we reported that zebrafish produces, through an alternative promoter, a novel truncated scl isoform, scl-b, which manifests a distinct spatiotemporal expression pattern from the previous reported full-length scl-a. Functional analysis reveals that scl-a and scl-b is redundant for the initiation of primitive hematopoiesis. However, they play different roles in primitive erythrocyte differentiation. In addition, the specification of definitive HSCs specifically requires scl-b but not scl-a. Furthermore, we provide evidence suggesting that the functional differences of scl-a and scl-b likely result from their different protein expression levels, possibly coming from their different protein stabilities. Taken together, our findings suggest that hematopoietic cells at different stage of hematopoietic hierarchy are properly governed by a gradient of Scl proteins established through the distinct spatial and temporal expression of these two different scl isoforms.Publication THE EFFECTS OF SUSPENDED SEDIMENTS ON PLANKTON(2018-08-23) SEW WEI XIN, GENEVIEVE; BIOLOGICAL SCIENCES; Peter Alan ToddIncreasing prevalence of anthropogenic activities (e.g. reclamation and dredging) in the coastal environment has resulted in elevated total suspended sediment (TSS) concentrations, impacting marine ecosystems. My thesis explores the impacts of these activities on the plankton community, beginning with a survey of the plankton and marine environment near two of the largest ongoing land reclamation projects in Singapore: Pulau Tekong and Tuas. I then experimentally investigated the impact of TSS on phytoplankton and zooplankton. Mesocosms were used to assess the effects of TSS concentration on a natural phytoplankton community. Using the copepod Acartia tonsa, TSS concentration, size and food availability were tested in combination to identify their effects on important biological functions. A. tonsa was also used to examine how TSS concentration and food availability affects swimming behaviour. My results shed light on the status of marine plankton in Singapore and act as a stepping-stone towards developing management tools for sediment pollution management.