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Chitinases have been implicated in aspects of growth and morphology of chitin containing fungi such as hyphal branching, anastamosis and, possibly, apical growth itself. Cloning of <i>Neurospora crassa</i> chitinase genes would allow a molecular genetic analysis of the involvement of chitinases in such phenomena through the study of regulation, subcellular location and the use of specific mutagenesis. The cytosolic chitinase was partially purified to a high specific activity and characterised. Allosamidin, the first specific inhibitor of chitinase to be described, was shown to be a potent inhibitor of the cytosolic chitinase. Direct sequencing of the chitinase protein was not possible due to the inability to purify the protein to homogeneity. However, protein sequence information was obtained from protein electroblotted onto Immobilon polyvinylide difluoride membrane, after first identifying the chitinase protein band <i>in situ</i> after polyacrylamide gel electrophoresis using a specific activity stain. Protein sequence data was used to design an oligonucleotide probe for the screening of a genomic library in the cosmid vector pSV50. Screening of the library resulted in the identification of a single positive cosmid. The region of cosmid 23D6 that showed hybridisation to the oligonucleotide was subcloned and sequenced. The oligonucleotide was shown to hybridise to a sequence that had a single mis-match. However, when the sequence surrounding the oligonucleotide binding site was converted to an amino acid sequence, using the reading frame for the chitinase protein sequence, no homology with the remaining chitinase protein sequence was seen.

Older adults improve in emotional well-being, and this may be a product of changes in motivation to regulate emotions or emotion regulation effectiveness. However, there are cognitive changes in old age that could make regulation harder in some contexts. The current set of studies sought to determine whether there were age-related improvements or deficits in ability to use distraction in two contexts. The first study examined use of distraction in a recovery context and found no age differences in emotional recovery when 1) using spontaneous self-distraction, 2) intentionally self-distracting, and 3) being distracted by another task. There was, however, evidence that the distracting task was the most effective way to recover from the negative induction. There was also some evidence that cognitive changes with age made it more difficult for older adults to limit negative thoughts in certain conditions. The second study contrasted use of distraction, positive reappraisal, and detached reappraisal by looking at success in terms of emotion regulation and impact on a subsequent cognitive task and later memory for the emotional stimuli. No age differences were found in emotion regulation success in this study either, but interesting differences in consequences of the three regulation strategies did emerge.

This paper investigates whether changes in internal control weakness (ICW) disclosure regulation affect the reliability of a firm’s disclosed ICW in a unique Canadian setting. In Canada, public firms have been required to provide internal control weakness disclosures since 2006. However, the credibility enhancement mechanisms (i.e., the implementation of effectiveness evaluation and CEO/CFO certification) were not adopted until 2008. Taking advantage of such an evolutionary process of regulations and inferring the reliability of the disclosed ICW from the magnitude of the negative association between disclosed ICW and investment efficiency, I first document that in the pre-adoption period, the association between Canadian firms’ disclosed ICW and their investment efficiency is insignificant; however, in the post-adoption period, the disclosed ICW negatively affects firms’ investment efficiency. This finding suggests that the credibility enhancement mechanisms improve the reliability of disclosed ICW in Canada. In addition, using the U.S. sample as a benchmark, I find that in the post-adoption period, the association is weaker between Canadian firms’ disclosed ICW and their investment efficiency, which is consistent with my prediction that the external audit requirement increases the reliability of the disclosed ICW. Overall, the study implies that changes in disclosure regulation lead to efficient resource allocation by improving the reliability of the information disclosed.

This paper investigates whether changes in internal control weakness (ICW) disclosure regulation affect the reliability of a firm’s disclosed ICW in a unique Canadian setting. In Canada, public firms have been required to provide internal control weakness disclosures since 2006. However, the credibility enhancement mechanisms (i.e., the implementation of effectiveness evaluation and CEO/CFO certification) were not adopted until 2008. Taking advantage of such an evolutionary process of regulations and inferring the reliability of the disclosed ICW from the magnitude of the negative association between disclosed ICW and investment efficiency, I first document that in the pre-adoption period, the association between Canadian firms’ disclosed ICW and their investment efficiency is insignificant; however, in the post-adoption period, the disclosed ICW negatively affects firms’ investment efficiency. This finding suggests that the credibility enhancement mechanisms improve the reliability of disclosed ICW in Canada. In addition, using the U.S. sample as a benchmark, I find that in the post-adoption period, the association is weaker between Canadian firms’ disclosed ICW and their investment efficiency, which is consistent with my prediction that the external audit requirement increases the reliability of the disclosed ICW. Overall, the study implies that changes in disclosure regulation lead to efficient resource allocation by improving the reliability of the information disclosed.

Metallothioneins (MTs) are low molecular weight, cysteine-rich, metal-binding proteins, which are proposed to have roles in essential trace metal homoeostasis and in the detoxification of metal ions. The genes encoding MTs have been isolated from a wide range of eukaryotes, although MT genes have not previously been isolated from prokaryotes. The polymerase chain reaction (PGR) was initially used to isolate a prokaryotic MT gene fragment from Synechococcus PCC 6301. PGR fragments were amplified using inosine-containing primers designed from the amino acid sequence of a prokaryotic MT. Subsequent cloning and nucleotide sequence analysis revealed that the deduced amino acid sequence of the PGR product corresponded to the amino acid sequence of the prokaryotic MT. The amplified product was thus part of the gene encoding the MT, and was designated smtA. The same primers used in the initial amplification were subsequently utilised for anchored PGR, to amplify the remainder of the coding region and the 3' and 5' flanking regions of the smtA gene. A genomic library was produced from Synechococcus PGG 7942 DNA and screened using the PGR products described above as probes. A genomic clone was isolated, nucleotide sequence analysis revealed the structure of the smt locus, two open reading frames, smtA and smtB, arranged in a divergent orientation about the smt operator/promoter region. The operator/promoter region contains the transcriptional and translational signals for the two genes and three regions that are candidate sites for interaction of regulatory proteins. The transcript start sites of the two genes were mapped within the operator/promoter region by primer extension analysis. An increase in the relative abundance of transcripts of both smt genes was studied in response to various metal ions in a series of northern blots. Inhibitor studies confirmed that the smtA gene is regulated at the transcriptional level. The 5' flanking region of the smtA gene conferred metal specific induction of the reporter gene lacZ. SmtB has sequence similarity to several prokaryotic regulatory proteins and contains a putative helix-turn-helix structural domain. Deletion analysis suggests that SmtB is a repressor of smtA expression. Subsequent work has confirmed that SmtB is a trans-acting repressor of expression from the smt operator/promoter.

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The trophoblast transcription factor glial cell missing-1 (GCM1) regulates asymmetric division of placental cytotrophoblast to form the differentiated syncytiotrophoblast. Reduced GCM1 expression is a key feature of the hypertensive disorder preeclampsia. In-silico techniques identified a novel calcium-dependent transcriptional repressor – DREAM as a regulatory candidate for GCM1. The overall objective of this thesis was to determine if DREAM regulates GCM1 expression and therefore villous trophoblast turnover. siRNA-mediated DREAM silencing in both BeWo cells and floating villous explants significantly upregulated GCM1 causing reduced cytotrophoblast proliferation. Calcium-dependency was demonstrated in both BeWo cells and floating villous explants by contrasting the effects of ionomycin and nimodipine. A direct interaction between DREAM and the GCM1 promoter was demonstrated using EMSA and ChIP assay. DREAM is a negative upstream regulator of GCM1 expression in human placenta that participates in calcium-dependent trophoblast differentiation.