Deregulated expression of circadian clock and clock-controlled cell cycle genes in chronic lymphocytic leukemia

Circadian rhythms are endogenous and self-sustained oscillations of multiple biological processes with approximately 24-h rhythmicity. Circadian genes and their protein products constitute the molecular components of the circadian oscillator that form positive/negative feedback loops and generate circadian rhythms. The circadian regulation extends from core clock genes to various clock-controlled genes that include various cell cycle genes. Aberrant expression of circadian clock genes, therefore, may lead to genomic instability and accelerated cellular proliferation potentially promoting carcinogenesis. The current study encompasses the investigation of simultaneous expression of four circadian clock genes (Bmal1, Clock, Per1 and Per2) and three clock-controlled cell cycle genes (Myc, Cyclin D1 and Wee1) at mRNA level and determination of serum melatonin levels in peripheral blood samples of 37 CLL (chronic lymphocytic leukemia) patients and equal number of age- and sex-matched healthy controls in order to indicate association between deregulated circadian clock and manifestation of CLL. Results showed significantly down-regulated expression of Bmal1, Per1, Per2 and Wee1 and significantly up-regulated expression of Myc and Cyclin D1 (P < 0.0001) in CLL patients as compared to healthy controls. When expression of these genes was compared between shift-workers and non-shift-workers within the CLL group, the expression was found more aberrant in shift-workers as compared to non-shift-workers. However, this difference was found statistically significant for Myc and Cyclin D1 only (P < 0.05). Serum melatonin levels were found significantly low (P < 0.0001) in CLL subjects as compared to healthy controls whereas melatonin levels were found still lower in shift-workers as compared to non-shift-workers within CLL group (P < 0.01). Our results suggest that aberrant expression of circadian clock genes can lead to aberrant expression of their downstream targets that are involved in cell proliferation and apoptosis and hence may result in manifestation of CLL. Moreover, shift-work and low melatonin levels may also contribute in etiology of CLL by further perturbing of circadian clock.     

Neuronal function of Tbx20 conserved from nematodes to vertebrates

The Tbx20 orthologue, mab-9, is required for development of the Caenorhabditis elegans hindgut, whereas several vertebrate Tbx20 genes promote heart development. Here we show that Tbx20 orthologues also have a role in motor neuron development that is conserved between invertebrates and vertebrates. mab-9 mutants exhibit guidance defects in dorsally projecting axons from motor neurons located in the ventral nerve cord. Danio rerio (Zebrafish) tbx20 morphants show defects in the migration patterns of motor neuron soma of the facial and trigeminal motor neuron groups. Human TBX20 is expressed in motor neurons in the developing hindbrain of human embryos and we show that human TBX20 can substitute for zebrafish tbx20 in promoting cranial motor neuron migration. mab-9 is also partially able to rescue the zebrafish migration defect, whereas other vertebrate T-box genes cannot. Conversely we show that the human TBX20 T-box domain can rescue motor neuron defects in C. elegans. These data suggest the functional equivalence of Tbx20 orthologues in regulating the development of specific motor neuron groups. We also demonstrate the functional equivalence of human and C. elegans Tbx20 T-box domains for regulating male tail development in the nematode even though these genes play highly diverged roles in organogenesis.     

“Occurrence of blaCTX-MGp1 and blaCTX-MGp26 in third generation cephalosporin-resistant and carbapenem- resistant bacterial isolates from southwest region of Saudi Arabia-a preliminary study”

This study was intended to identify the genes responsible for ESBL- and carbapenemase-producing bacterial isolates obtained from Jizan region. A hospital-based cross-sectional study was conducted over a period of 3 months (15th November 2018–15th February 2019). Fifty non-duplicate, 3rd-generation cephalosporin and carbapenem-resistant isolates were collected from microbiology lab of a tertiary care hospital in Jizan province and were screened for ESBLs and MBLs by phenotypic methods (CDT). The positive isolates (by phenotypic method) were then scanned for the presence of bla_ESBLs and bla_NDM-₁ genes, respectively, by PCR.As a result, 10% isolates showed imipenem-cephalosporin co-resistance whereas 92% (46/50) of isolates were found to be ESBL producers by CDT. The maximum occurrence was observed for bla_CTX-M (70%), followed by bla_SHV (16%) and least occurrence was noted for bla_TEM (12%). Moreover, 97% isolates (34/35) were of bla_CTX-MGroup1 but one isolate showed the presence of bla_CTX-M Group26. Despite the co-resistance of cephalosporin and carbapenem, 14% (7/50) were found to be MBL producer on phenotypic detection by Combination Disc Test (CDT), whereas all the isolates were found to be negative for bla_NDM-1. Hence bla_CTX-MGroup1 is present in quite high fraction followed by bla_SHV in the bacterial isolates of Jizan region. Moreover, the occurrence of bla_CTX-M Group1 and bla_CTX-M Group26 in clinical isolates from the Jizan region of Saudi Arabia has been reported for the first time.     

Effect of sequential deletion of extra N-terminal residues on the structure and stability of yeast iso-1-cytochrome-c

A sequence alignment of yeast cytochrome-c (y-cyt-c) with mammalian cyts-c shows that the yeast protein has a five residue long N-terminal extension. A question arises: Does this N-terminal extension play any roles in the stability, structure, and folding of the yeast protein? To answer this question, in silico and in vitro studies were carried out on the wild type (WT) protein and its five deletants (Δ(?5/?5), Δ(?5/?4), Δ(?5/?3), Δ(?5/?2), and Δ(?5/?1) where Δ denotes the deletion and the numbers refer to the residues deleted, e.g. Δ(?5/?1) denotes the deletion of residues numbered from ?5 to ?1 (TEFKA), while Δ(?5/?2) denotes the deletion of resides numbered from ?5 to ?2 (TEFK) and so on). The main conclusion of the in silico study is that the order of stability of deletants and WT protein is Δ(?5/?4) > WT > Δ(?5/?3) > Δ(?5/?5) > Δ(?5/?1) ~ Δ(?5/?2). In vitro studies involved (i) measurements of thermodynamic stability of all proteins by differential scanning calorimetry and from sigmoidal curves of two different structural properties ([θ]₂₂₂, a probe for detecting change in secondary structure, and Δε₄₀₅, a probe for detecting alteration in the heme environment), and (ii) characterization of all proteins by various spectral properties. The main conclusions of the in vitro studies are as follows: (i) The order of thermodynamic stability of all proteins is in excellent agreement with that predicted by in silico studies, and (ii) A sequential deletion of the N-terminal extension has no effects on protein structure and folding.     

Monoamine oxidase in amphistomes and its role in worm motility

The quantitative assay of mitochondrial monoamine oxidase (MAO) activity revealed a higher enzyme level in Explanatum explanatum than Gastrothylax crumenifer. The specific MAO inhibitors, chlorgyline, pargyline, deprenyl and nialamide produced different degrees of interspecific inhibition. The differential effects on enzyme activity of chlorgyline and deprenyl suggests the possible existence of polymorphic forms of the enzyme, MAO-A and MAO-B, in amphistomes. These specific inhibitors also had a differential influence on the in vitro motility of amphistomes, further indicating the involvement of different forms of MAO in the oxidative deamination of biogenic monoamines which might be partly responsible for neuromuscular coordination in amphistomes. The experimental procedures used in this study could be conveniently used for quick screening and evaluation of some of the qualitative effects of anthelmintic drugs under in vitro conditions.     

Preliminary studies in numerical systematics of the Paramphistomoidea (Digenea) from domestic Artiodactyla of Northern India

The systematic relationships between nine taxa (species) of paramphistomes belonging to the families Cladorchiidae, Gastrodiscidae, Gastrothylacidae and Paramphistomidae from domestic buffaloes Bubalus bubalis and pigs Sus scrofa were examined using SEM revealed two types of papillae occurring commonly on the oral cone and less frequently around the gonopore and acetabulum. Non-pitted papillae occur on the oral cone in Paramphistomum epiclitum, Calicophoron calicophorum, Explanatum (=Gigantocotyle) explanatum, Orthocoelium scoliocoelium, Gastrothylax crumenifer and Olveria indica. These papillae are also evident around the gonopore in Calicophoron papillosum, O. scoliocoelium and O. indica, and the ventral pouch opening in G. crumenifer. Ciliated papillae occur on the oral cone in C. papillosum and Fischoederius elongatus. Morphological and immunological observations were employed to classify paramphistomes using phenetic and phylogenetic approaches to systematics. Classifications based on phenetic methods compared taxa according to overall similarity of their characteristics and produced largely discordant results. Classifications based on phylogenetic methods compared taxa according to the genealogical descent of their characteristics and produced largely concordant results. These are in agreement with the widely accepted scheme of classification proposed by Stiles & Goldberger (1910) in which Gastrothylacidae and Paramphistomidae occur as natural groups. Furthermore, the Gastrothylacidae appear to share common ancestry with a member of the Paramphistomidae and are consequently nested within the latter group.This study forms part of the requirements for a thesis submitted by the first author to Q.U.B. for the degree of Doctor of Philosophy.This study forms part of the requirements for a thesis submitted by the first author to Q.U.B. for the degree of Doctor of Philosophy.     

Quantitative studies on acetylcholinesterase in seven species of digenetic trematodes

Quantitative estimation of absolute levels and in vitro release of acetylcholinesterase (AChE) in seven species of digenetic trematodes: Isoparorchis hypselobagri from the swim bladder of catfish, Wallago attu; Srivastavaia indica and Gastrothylax crumenifer from the rumen, and Gigantocotyle explanatum from the liver of the water buffalo, Bubalus bubalis; Fasciolopsis buski, Echinostoma malayanum from the small intestine and Gastrodiscoides hominis from the caecum of the pig, Sus scrofa revealed that the enzyme is present in remarkably high quantities in species which inhibit gastrointestinal tract compared with those that parasitize liver and swim bladder. The rate of in vitro release of AChE also varies with the species which supports the view that such differential secretion probably takes place in situ as well to counteract peristalsis and it is a biochemical adaptation on the part of these trematodes.