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Biomedical subjects

T Jowett

Publications and source records attributed to T Jowett.

At least 37 records · Page 2Linked to original sources
PubMed1993

Cloning of the zebrafish krox-20 gene (krx-20) and its expression during hindbrain development.

To begin to examine the function of genes that control early development in the hindbrain, we have screened an embryonic zebrafish cDNA library with a murine krox-20 gene probe that contained the conserved zinc finger regions. We have isolated two overlapping cDNAs, zf187 and zf201 which are homologues of the murine krox-20 gene. The N-terminal of the longest cDNA (zf201) contains two acidic regions identical to those of the murine krox-20. This indicates that the functional organisation of these proteins is probably conserved. Northern Blot analysis identified a single transcript of 2.0 kb. Wholemount in situ hybridisation established that expression of the zebrafish gene (krx-20) first appears at 100% epiboly as a single anterior domain of the prospective neuroepithelium, followed very soon after by a second more posterior domain. The alternating pattern of expression of this gene in rhombomeres(r) r3 and r5 is apparent by 12 hr post-fertilisation, that is prior to the morphological appearance of the rhombomeres. Around 14 hr neural crest migration begins from the dorsal surface of r5, moving caudally into r6 and then ventrally towards the pharyngeal arches. Crest migration is not apparent at or after 16 hr. No neural crest migration was observed from r3. Expression of krx-20 is down regulated firstly in r3 around 26 hr and later in r5 around 30 hr.

Amino Acid Sequence↗
PubMed1991

Cloning of fish zinc-finger genes related to Krox-20 and Krox-24.

Southern hybridization suggests that the zebrafish genome contains multiple zinc-finger genes related to the putative mouse developmental genes, Krox-20 and Krox-24. The polymerase chain reaction was employed to amplify and clone the zinc-finger regions of genes related to Krox-20, from two fish species and, for comparison, mouse, hamster and fox. DNA sequence analyses suggest that the genes cloned include the guppy homologue of Krox-20 and the zebrafish homologue of Krox-24, and that these genes diverged prior to the separation of the lineages leading to teleosts and to mammals.

Amino Acid Sequence↗
PubMed1991

The major serum protein of Drosophila larvae, larval serum protein 1, is dispensable.

Null alleles of all three genes (alpha, beta and gamma) coding for the major serum protein of Drosophila larvae, larval serum protein 1, have been characterized at the protein level and by analysis of their RNA and DNA. Each allele completely lacks one polypeptide chain. beta shows an unaltered gene but reduced levels of RNA, alpha shows restriction-fragment-length polymorphism and shows low levels of transcript, while gamma is a deletion. The three null alleles have been combined to give a strain without the gene for larval serum protein 1 which survives.

Alleles↗
PubMed1991

Mammalian genes expressed in Drosophila: a transgenic model for the study of mechanisms of chemical mutagenesis and metabolism.

Mammalian cytochrome P450s provide our first line of defence against the toxic effects of environmental chemicals. Ironically these enzymes also convert some compounds to their ultimate toxic or mutagenic species. Our knowledge of these mammalian enzymes and the role they play in chemical toxicity and mutagenesis has stemmed mostly from in vitro studies. In order to establish the role of specific enzymes in the toxicological response in vivo we have generated transgenic Drosophila which express mammalian cytochrome CYP2B1, which is a member of a large gene family encoding several important drug metabolising enzymes. The gene was fused to a Drosophila promoter which confers expression in the larval fat body. Using the Somatic Mutation And Recombination Test (SMART) we have demonstrated that transgenic larvae expressing the P450 are hypersensitive to the anticancer drug cyclophosphamide, a procarcinogenic substrate which is activated by the enzyme. This work demonstrates the potential of such transgenic Drosophila strains as an in vivo model for studying the role of specific mammalian drug metabolising enzymes in the pathways and metabolic cascades associated with the action of cytotoxic and carcinogenic chemicals, and also the chemical properties of specific classes of mutagen to be determined.

Animals↗
PubMed1990

Specific binding sites for 125I-labelled human luteinizing hormone (hLH) in microsomal fractions from Drosophila.

Microsomal fractions prepared from Drosophila flies and larvae bound 125I-labelled human luteinizing hormone (hLH) in a displaceable manner. Binding increased linearly with increasing microsome concentration, and was dependent on the pH and metal ion concentration of the medium, and on the temperature and duration of incubation. Scatchard plots of 125I-hLH binding demonstrated the presence of two distinct binding sites, one with high affinity and low capacity, the other with low affinity and high capacity. 125I-hLH binding was inhibited in a dose-dependent fashion by partially purified human gonadotropin preparations, but not by other proteins, hormones and peptides. Highly purified human chorionic gonadotropin (hCG) or hLH preparations were much less effective at inhibiting 125I-hLH binding to Drosophila microsomes than partially purified gonadotropins. This was due to the presence of a contaminant present in partially purified commercial hCG preparations. Chromatography of crude hCG preparations clearly resolved the LH-binding inhibitor from hCG. Moreover, whereas hCG (and epidermal growth factor-like factors present in crude hCG preparations) were not bound to CM-Sepharose, the Drosophila LH-binding inhibitor was adsorbed strongly. Fractionation of 125I-hLH tracer on concanavalin A-Sepharose suggested that Drosophila binding sites did not preferentially bind a subfraction of the hormone tracer. These data suggest that high affinity binding sites for hLH/hCG-like molecules are present in Drosophila microsomes.

Animals↗
PubMed1989

Maturation of thyroid function in normal human foetuses.

The functional maturation of an independent foetal thyroid activity was investigated in the present study. Serum concentrations of total T4, free T4, TSH and TBG were measured in 23 foetuses between 18 and 31 weeks' gestational age. Foetal samples were collected by transabdominal needling from the placental cord insertion. TT4, FT4, TBG and TSH levels significantly increased with gestational age. FT4 levels were comparable with the adult range by 28 weeks' gestation; TBG levels reached adult values at approximately 30 weeks, while TT4 was lower than adult levels throughout the whole period studied. TSH values were, in all cases, higher than the normal adult range. A significant positive correlation was present between TT4 and TBG, TT4 and TSH, and TBG and TSH levels; on the contrary, no correlation was demonstrated between FT4 and TSH levels. The TSH/TT4 ratio significantly decreased with gestational age. The results suggest an incomplete responsiveness of the foetal thyroid gland to TSH, while the feedback control system between pituitary and thyroid is operating at a different set point from that in post-natal life. The normal range of thyroid parameters established is of clinical relevance for the antenatal diagnosis, and eventual treatment of thyroid disorders that may seriously damage foetal development and maturation.

Fetus↗
PubMed1988

A novel human cytochrome P450 gene (P450IIB): chromosomal localization and evidence for alternative splicing.

We have isolated from a single human liver cDNA library two clones which are highly homologous (78% over the coding region) to the major phenobarbital-inducible P450 from rat (P450IIB1). This is the first direct demonstration of the presence of the P450IIB gene subfamily in humans. This subfamily is much less extensive than the rodent homologues, but does appear to contain at least two genes. Of the cDNA clones isolated one is apparently normally spliced, whereas the other lacks exon 8 and retains all or part of intron 5. Both clones contain transcribed Alu sequences. The human P450IIB gene has been located to chromosome 19q12----19q13.2 using a probe derived from intron 5, and is close to the CYP 2A locus encoding cytochrome P450IIA2. Restriction fragment length polymorphisms have been found with the enzymes BamHI and MspI which will enable linkage to be determined between these two loci.

Amino Acid Sequence↗
PubMed1986

Hepatitis A virus replication in tamarins and host immune response in relation to pathogenesis of liver cell damage.

Hepatitis A virus (HAV) shedding in the faeces, appearance of HAV-Ag (antigen) in the liver, and development of humoral immunity to HAV have been studied in experimentally infected tamarins. The appearance of liver damage measured by transaminase elevation and histology, in relation to the above variables, suggests that the virus is not cytopathic and the immune system contributes to the production of liver cell damage. Preliminary data suggest that HAV replication may occur in the mucosa of the small intestine and in the liver.

Animals↗
PubMed1986

Regulation of synthesis of larval serum proteins after transplantation of larval fat body into adult Drosophila melanogaster.

The larval serum proteins, LSP1 and LSP2, of Drosophila melanogaster are synthesized by the fat body during the third instar. We examined the potential for LSP synthesis by fat body implants in adult flies. Fat body from third instar donors will continue to synthesize LSPs in both males and females. Implants from late second instar larvae will start synthesizing LSP1 and LSP2 in females but only LSP1 in males, suggesting that regulation of these proteins is not the same and that the physiological milieu in the two sexes differs. The newly synthesized LSPs are secreted into the hemolymph for approximately 48 hr when secretion stops but synthesis continues. This sequence follows the pattern for LSP secretion in situ. Fat body from mid second instar larvae is variable in its ability to synthesize LSPs. LSPs are not detected in implants from first instar larvae despite there being a high level of protein synthesis in the implant and considerable growth of the fat body cells. We conclude that there is a critical stage of differentiation during the latter half of the second instar when the fat body becomes independent of the larval milieu and can synthesize LSPs in the adult.

Adipose Tissue↗
PubMed1985

The regulatory domain of a larval serum protein gene in Drosophila melanogaster.

DNAse I hypersensitive sites have been shown to be present at the 5' end of each of the larval serum protein genes (LSPs) when they are expressed in the fat body. They are absent from the same inactive genes in embryos. A 4.4-kb genomic fragment containing the coding region for LSP 1 alpha and 1.5 kb of 5'-flanking sequence has been reintroduced into the germ line of a strain containing an electrophoretic variant form of the alpha gene. The expression of the reintegrated gene is normal with respect to the amount of protein produced and the temporal and spatial distribution of the transcript. A comparison has been made of the chromatin configuration of the reintroduced and endogenous alpha genes and the same pattern of DNase I hypersensitivity is found. The hypersensitive regions have been located on the 5' sequence of the LSP 1 alpha gene and their sequences compared with known hypersensitive regions of other genes.

Journal Article↗