Biomedical subjects

Lee Zhang

Publications and source records attributed to Lee Zhang.

8 recordsLinked to original sources

PubMed2006

Kdr allelic variation in pyrethroid resistant mosquitoes, Culex quinquefasciatus (S.).

The loss of target site sensitivity to insecticides resulting from a substitution of leucine to phenylalanine, termed the kdr mutation, in the voltage-gated sodium channel of the insect nervous system is known to be important in insecticide resistance. However, little is known about the molecular basis of the genotype and kdr-mediated resistance phenotype relationship. This study investigated whether the functional polymorphism of the L-to-F kdr mutation that determines resistance phenotype undergoes DNA variation or goes through transcriptional regulatory variation. We detected no correlation for the kdr allele at the genomic DNA level with levels of susceptibility and resistance to insecticide. However, we find a strong correlation between the kdr allelic expression and levels of insecticide resistance and susceptibility through RNA allelic variation and RNA editing. These findings shed new light on the role of transcriptional regulation in the kdr-mediated resistance in mosquitoes and its connection with the genotype-resistance phenotype relationship.

Alleles↗

PubMed2006

Sodium channel gene expression associated with pyrethroid resistant house flies and German cockroaches.

The voltage-gated sodium channels of the insect nervous system are the primary target of DDT and pyrethroid insecticides. The loss of target site sensitivity to insecticides resulting from a substitution of leucine to phenylalanine, termed the L-to-F kdr mutation, in the sodium channel of the insect nervous system is known to be important in insecticide resistance. Yet, little is known about the molecular basis underlying the genotype and kdr-mediated resistance phenotype relationship. Here we report a systematic study of resistance-associated kdr allelic expression within and among resistant and susceptible house fly and German cockroach populations. We compared genomic DNA and RNA sequences within the same individuals from different insect strains, finding no correlation for the kdr allele at the genomic DNA level with levels of susceptibility or resistance to insecticide. However, there was a strong correlation between kdr allele expression and the levels of insecticide resistance. This correlation is probably regulated through RNA variation and RNA editing. These results suggest a role for posttranscriptional regulation in the connection of the sodium channel genotype and its mutation-mediated resistance phenotype.

Alleles↗

PubMed2005

Resistance in the mosquito, Culex quinquefasciatus, and possible mechanisms for resistance.

Two mosquito strains of Culex quinquefasciatus (Say), MAmCq(G0) and HAmCq(G0), were collected from Mobile and Huntsville, Alabama, respectively. MAmCq(G0) and HAmCq(G0) were further selected in the laboratory with permethrin for one and three generations, respectively. The levels of resistance to permethrin in MAmCq(G1) (after one-generation selection) and HAmCq(G3) (after three-generation selection) increased rapidly. Resistance to permethrin in MAmCq(G1) and HAmCq(G3) was partially suppressed by piperonyl butoxide (PBO), S,S,S-tributylphosphorotrithioate (DEF) and diethyl maleate (DEM), inhibitors of cytochrome P450 monooxygenases, hydrolases and glutathione S-transferases (GST), respectively, suggesting these three enzyme families are important in conferring permethrin resistance in both strains. A substitution of leucine to phenylalanine (L to F) resulting from a single nucleotide polymorphism (SNP), termed the kdr mutation, in the para-homologous sodium channel gene has been reported as a very common mutation associated with pyrethroid resistance of insects. A 341-bp sodium channel gene fragment, where the kdr mutation resides, was generated by PCR from genomic DNAs of Cx. quinquefasciatus strains. We found that the kdr mutation was present in both permethrin-selected and unselected HAmCq and MAmCq mosquito populations, suggesting that the kdr mutation plays the role in permethrin resistance. There was no significant change in the frequency and heterozygosity of the A to T SNP for the kdr allele between permethrin-selected and unselected MAmCq and HAmCq mosquitoes, indicating that other mechanisms are involved in the evolution of resistance in mosquitoes selected by permethrin in the laboratory.

Animals↗

PubMed2005

Chlorpyrifos resistance in mosquito Culex quinquefasciatus.

Two mosquito strains of Culex quinquefasciatus Say, MAmCq and HAmCq, were collected from Mobile and Huntsville, AL, respectively, after the control of mosquitoes with insecticides proved difficult. A synergism study showed that resistance to chlorpyrifos in MAmCq and HAmCq was not suppressed by piperonyl butoxide (PBO) and S,S,S,-tributylphosphorotrithioate (DEF), suggesting that P450 monooxygenase- and hydrolase-mediated detoxication does not contribute to chlorpyrifos resistance in either strain. Diethyl maleate (DEM) did not cause any significant change in the level of chlorpyrifos toxicity to HAmCq. However, DEM enhanced toxicity of chlorpyrifos to MAmCq 2.5-fold, indicating that glutathione S-transferase (GST)-mediated detoxication may play a minor role in the resistance of MAmCq. An inhibition study of acetylcholinesterase (AChE) by chlorpyrifos showed that bimolecular rate constants (Ki) of chlorpyrifos for the inhibition of AChE in adults and larvae of the susceptible S-Lab strain were 2.2- and 1.9-fold higher, respectively, than in the HAmCq strain and 3.4- and 3.8-fold higher than in the MAmCq strain. The single mutation, G119S, resulting from a single nucleotide polymorphism (SNP), G to A, in ace-1 acetylcholinesterase gene was present in HAmCq and MAmCq mosquitoes. The frequency of the heterozygote for the G119S mutant allele in the HAmCq and MAmCq mosquito populations was 0.25 and 0.45, respectively, and no individuals in either of these mosquito strains were homozygous for the A allele. It thus seems likely that the presence of heterozygous individuals for the G119S allele in HAmCq and MAmCq populations may be a response to the insensitivity of AChE observed in these two mosquito strains.

Acetylcholinesterase↗

PubMed2004

CYP4AB1, CYP4AB2, and Gp-9 gene overexpression associated with workers of the red imported fire ant, Solenopsis invicta Buren.

Two cytochrome P450 genes, CYP4AB1 and CYP4AB2, and the Gp-9 gene were identified as being specifically overexpressed in workers of the red imported fire ant using PCR-selected subtractive hybridization and cDNA array techniques. Full-length CYP4AB1 and CYP4AB2 were cloned and sequenced. The cDNA sequences of CYP4AB1 and CYP4AB2 have open reading frames of 1389 and 1533 nucleotides encoding proteins of 463 and 511 amino acid residues, respectively. Northern blot analysis was performed to compare expression levels of CYP4AB1, CYP4AB2, and Gp-9 for different developmental stages and castes of fire ants. We demonstrate that the expression of these three genes is developmentally and caste specifically regulated in red imported fire ants. Levels of CYP4AB1 mRNA were undetectable in 3rd+4th instars, worker pupae, and alate (mixed sex) pupae; readily detectable in male and female alates; increased (approximately 3-fold) in the queens; and rose to a maximum (13-fold) in workers. Similarly, the expression of CYP4AB2 mRNA was undetectable in 3rd+4th instars, worker pupae, and alate pupae; low in male and female alates and queens; and increased (approximately 7-fold) in workers. Levels of Gp-9 mRNA were readily detectable in male alates; increased (approximately 3-fold) in female alates; and reached a maximum (approximately 12-fold) in workers. Their caste-specific overexpression suggests the functional importance of CYP4AB1, CYP4AB2, and Gp-9 in workers of the red imported fire ant.

Amino Acid Sequence↗

PubMed2003

Overexpression of CYP4G19 associated with a pyrethroid-resistant strain of the German cockroach, Blattella germanica (L.).

A new cytochrome P450 gene, CYP4G19, was identified and isolated as a differentially expressed gene between insecticide susceptible ACY and resistant Apyr-R German cockroach strains using PCR-selected subtractive hybridization and cDNA array techniques. The cDNA sequence of CYP4G19 has an open ready frame of 1638 nucleotides encoding a putative protein of 546 amino acid residues. Sequence analysis shows that CYP4G19 putative protein contains (1) a highly hydrophobic N terminus, (2) a P450 protein signature motif, FXXGXRXCXG, known to be an important ligand for heme binding, and (3) an important characteristic motif, EVDTFMFEGHDTT, for the family 4. Northern blot analysis indicated that levels of CYP4G19 expression were low in eggs, nymphs, and adults of the susceptible ACY strain with a similar expression pattern. The expression of CYP4G19 in the resistant Apyr-R strain was developmentally regulated, with very low expression in eggs, increasing in nymphs, and reaching a maximum in both female and male adults. Comparison of CYP4G19 expression between ACY and Apyr-R strains indicated that there was no difference in their eggs, but expression was higher ( approximately 1.7-fold) in the nymphs and much higher ( approximately 5-fold) in the male and female adults of the Apyr-R strain. Levels of CYP4G19 mRNA were readily detectable in head+thorax tissues and increased ( approximately 5-fold) in the abdomens of the ACY strain. In the Apyr-R strain, however, levels of CYP4G19 mRNA were relatively low in head+thorax tissues and were about 7-fold increase in the abdomen. Although expression patterns of CYP4G19 in head+thorax and abdomen tissues were similar (i.e. lower in head+thorax tissues and higher in abdomen tissues) in both the ACY and Apyr-R strains, the overexpression was more evident in the Apyr-R strain in both head+thorax and abdomen tissues than in the ACY strain.

Amino Acid Sequence↗

PubMed2002

A pH sensor based on force generated by pH-dependent polymer swelling.

We have demonstrated the feasibility of a new type of pH sensor by combining a bead of porous lightly crosslinked diethanolamine derivatized poly(vinylbenzyl chloride) with a strain gauge, i.e. a pressure sensitive resistor. The polymer bead is toughened with Kraton G1652, a styrene-ethylene,butylene-styrene triblock copolymer. The sensor is constructed so that the shrunken form of the bead is held in contact with the strain gauge with a small force. Increases in the hydrogen ion concentration protonate the diethanolamine introducing a positive charge onto the polymer backbone. This results in an electrostatic swelling force that causes the polymer to swell. This is detected as a change in strain gauge resistance that is readout via a Wheatstone bridge. When the pH of 0.10 M buffers is changed from 10 to 4, the response time is 390 s for a bead that is 0.25 mm in diameter in the shrunken state. The response varies with the square of the bead radius. The magnitude of the response is highly correlated with the penetration modulus, a measure of the extent to which the bead resists deformation when subjected to an external force. The response to pH appears to be shifted by the application of pressure in the sensor. This instrumentally simple approach to sensing has the potential to be stable and long-lived if the polymer bead can undergo a large number of swelling/shrinking cycles without changing mechanical properties.

Journal Article↗

PubMed2002

Identification of two new cytochrome P450 genes and their 5'-flanking regions from the housefly, Musca domestica.

Two new cytochrome P450 cDNAs, named CYP28B1 and CYP4G13v2, and their 5'-flanking regions were cloned and sequenced from a housefly strain, ALHF. The cDNA sequences of CYP28B1 and CYP4G13v2 have open reading frames of 1449 and 1653 nucleotides encoding proteins of 483 and 551 amino acid residues, respectively. Sequence analysis shows that both CYP28B1 and CYP4G13v2 putative P450 proteins contain: (1) a highly hydrophobic N terminus; (2) a P450 protein signature motif, FXXGXRXCXG, known as the important ligand for heme binding; (3) a motif, YXXAXXXEXXR, which is a conserved P450 sequence coinciding with Helix K; and (4) a typical aromatic sequence, A(1)XXPXXA(2)XPXBA(3), which is conserved within most P450s. The 5'-flanking regions of CYP28B1 (>2kb) and CYP4G13v2 (>1 kb) were isolated from adaptor-ligated ALHF genomic DNA libraries. The transcription start points of CYP28B1 and CYP4G13v2 were mapped to 176 and 163 nucleotides upstream of the ATG translation start codon within the conserved arthropod promoter elements of TCATT and ACAGT, respectively. Possible regulatory binding sites for general transcription factors, Sp1 and AP1, were mapped in the 5' promoter regions of CYP28B1 whereas TFIID and Oct-1 were mapped in CYP4G13v2. Five conserved cis-acting elements for tissue- or cell-specific transcription regulatory factors were identified in the promoter regions of both P450 genes. A structure of five 153-nucleotide (nt) highly identical repeats and two partial repeat sequences were found in the promoter region of CYP28B1. The homologous (90% identity) sequences of the 153-nt repeat were also found in the promoter region of CYP4G13v2. The homologous sequences of the repeat in other insect P450 gene promoter regions are discussed.

5' Flanking Region↗