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A R Place

Publications and source records attributed to A R Place.

At least 19 recordsLinked to original sources
PubMed2005

Sex change strategy and the aromatase genes.

Sequential hermaphroditism is a common reproductive strategy in many teleosts. Steroid production is known to mediate both the natural and induced sex change, yet beyond this the physiology directing this process has received little attention. Cytochrome P450 aromatase is a key enzyme in the hormonal pathway catalysing the conversion of sex steroids, androgens to oestrogens, and thus is highly relevant to the process of sex change. This study reports the isolation of cDNA sequences for aromatase isoforms CYP19A1 and CYP19A2 from teleost species representing three forms of sexual hermaphroditism: Lates calcarifer (protandry), Cromileptes altivelis (protogyny), and Gobiodon histrio (bi-directional). Deduced amino acid analysis of these isoforms with other reported isoforms from gonochoristic (single sex) teleosts revealed 56-95% identity within the same isoform while only 48-65% identity between isoforms irrespective of species and sexual strategy. Phylogenetic analysis supported this result separating sequences into isoform exclusive clades in spite of species apparent evolutionary distance. Furthermore, this study isolates 5' flanking regions of all above genes and describes putative cis-acting elements therein. Elements identified include steroidogenic factor 1 binding site (SF-1), oestrogen response element (ERE), progesterone response element (PRE), androgen response element (ARE), glucocorticoid response elements (GRE), peroxisome proliferator-activated receptor alpha/retinoid X receptor alpha heterodimer responsive element (PPARalpha/RXRalpha), nuclear factor kappabeta (NF-kappabeta), SOX 5, SOX 9, and Wilms tumor suppressor (WTI). A hypothetical in vivo model was constructed for both isoforms highlighting potential roles of these putative cis-acting elements with reference to normal function and sexual hermaphroditism.

Animals↗
PubMed2001

Expression of P450(arom) in Malaclemys terrapin and Chelydra serpentina: a tale of two sites.

The formation of estrogens from androgens in all vertebrates is catalyzed by the "aromatase" complex, which consists of a membrane bound P(450) enzyme, P(450) aromatase (which binds the androgen substrate and inserts an oxygen into the molecule), and a flavoprotein (NADPH-cytochrome P450 reductase). Among vertebrates, the two major sites of aromatase expression are the brain and gonads. Given the importance of estrogen in reptile sex determination, we set out to examine whether P450arom was involved in the initiation and/or stabilization of sex determination in turtles. We examined the expression of aromatase activity in the brain and gonads of two turtle species exhibiting temperature dependent sex determination (TSD), the diamondback terrapin (Malaclemys terrapin), and the common snapping turtle (Chelydra serpentina). Estradiol when applied at stage 14 of the terrapin induces expression of aromatase in the gonad of embryos incubated at male temperatures (26.5 degrees C). The level of expression is similar to that of a normal embryonic ovary. When applied at stage 22, estradiol does not induce aromatase expression in the terrapin. The xenoestrogen, nonylphenol, sex reverses terrapin embryos at 26.5 degrees C. Letrazole, a nonsteroidal aromatase inhibitor, suppresses aromatase activity in the brain at either incubation temperature. Ovotestes are produced by letrazole administration in the terrapin when incubated at 30.5 degrees C. In the snapping turtle at stage 23, gonadal and brain aromatase activity in embryos incubated at female temperatures (30.5 degrees C) is nearly half that exhibited in terrapin embryos at the same temperature. Moreover, letrazole administration suppresses aromatase expression to nearly basal levels. At male incubation temperatures (26.5 degrees ), brain aromatase expression is nearly three times higher than at female temperatures, while gonadal expression levels are nearly one third lower. However, the gonadal expression levels at male temperatures in the snapping turtle are nearly 25 times higher than that found in the terrapin. Estradiol administration elevates this level nearly three fold. These data suggest that is not merely the expression of aromatase that is important for ovarian development, but that the level of expression may be more important.

Animals↗
PubMed2001

The 5'-flanking regions of CYP19A1 and CYP19A2 in zebrafish.

This report describes the structure of the 5'-flanking regions of both the CYP19A1 and A2 genes that were isolated from the genome of the zebrafish (Danio rerio). Consensus sequences of three cAMP-responsive elements (CRE), an aryl hydrocarbon-responsive element (AhR/Arnt), a steroidogenic factor 1 (SF-1) site, and a TATA box were observed in the 5'-flanking region of CYP19A1. In contrast, the 5'-flanking region of CYP19A2 was located upstream of an untranslated exon and possessed consensus sequences of a single CRE, an estrogen-responsive element (ERE), a peroxisome proliferator-activated receptor alpha/retinoid X receptor alpha heterodimer-responsive element (PPARalpha/RXRalpha), and a TATA box. Primer extension analysis revealed that the predominant transcription initiation sites for CYP19A1 and A2 transcripts were 28 and 91 bp upstream from the putative translation initiation codon, respectively. These analyses indicate that substantially different regulators, including a variety of environmental xenobiotics, control the expression the two CYP19 genes.

5' Untranslated Regions↗
PubMed2001

Alligator aromatase cDNA sequence and its expression in embryos at male and female incubation temperatures.

In all species of crocodilians, sex is determined not by genetic mechanisms, but by the temperature at which the egg is incubated. In the American alligator (Alligator mississippiensis) the thermosensitive period (TSP) for sex determination is a 7- to 10-day window within stages 21-24 of development, around the middle third of the incubation period. Treating embryos with estrogen during the TSP produces female offspring, even at male incubation temperatures. Conversely, blocking embryonic estrogen synthesis at female-inducing temperature prevents development of the female phenotype. Therefore, it has been suggested that estrogen plays a role in determination of sex in the alligator. Estrogen is produced from an androgen substrate by cytochrome P450 aromatase (CYP19). If estrogen plays a critical role in sex determination, there should be differences in aromatase expression between embryos at male- and female-producing temperatures during the TSP. Therefore, to address this question, we cloned and characterized the alligator CYP19 cDNA. Based on the sequence information, a quantitative kinetic reverse transcriptase-polymerase chain reaction (TaqMan) assay was designed to measure expression of the alligator aromatase gene in RNA extracted from the gonadal and brain regions of alligator embryos incubated at male- or female-producing temperatures from prior to the TSP through hatching. Aromatase expression was detected in the brain region from the earliest stage tested (stage 20) through hatching. The hypothalamus had significantly higher expression than the forebrain or hindbrain in both male and female embryos. Expression was not significantly different in the gonadal region between embryos at male and female temperatures until after the TSP, when there was a dramatic increase in expression at female temperature. These data indicate that aromatase expression and, thus, estrogen production, are not the initial trigger for sex determination but play an essential role in ovarian differentiation in the alligator. J. Exp. Zool. 290:439-448, 2001.

Alligators and Crocodiles↗
PubMed2001

Developmental expression of cytochrome P450 aromatase genes (CYP19a and CYP19b) in zebrafish fry (Danio rerio).

Cytochrome P450 aromatase (CYP19) is the terminal enzyme in the steroidogenic pathway that converts androgens (e.g., testosterone) into estrogens (e.g., estradiol). Regulation of this gene dictates the ratio of androgens to estrogens; therefore, appropriate expression of this enzyme is critical for reproduction as well as being pivotal in sex differentiation for most vertebrates. It is assumed that most vertebrates have a single CYP19 gene that is regulated by multiple tissue-specific promoter regions. However, the zebrafish (Danio rerio) has two genes (CYP19a and CYP19b), each encoding a significantly different protein and possessing its own regulatory mechanism. The primary purpose of this study was to determine the pattern of expression of each of the CYP19 genes in the developing zebrafish. A fluorescent-based method of real-time, quantitative RT-PCR provided the sensitivity and specificity to determine transcript abundance in single embryos/juveniles harvested at days 0 through 41 days post-fertilization (dpf), which encompasses the developmental events of sex determination and gonadal differentiation. CYP19 transcripts could be detected as early as 3 or 4 dpf, (CYP19a and CYP19b, respectively) and peak abundance was detected on day five. In general, the CYP19 genes differed significantly in the ontogeny of their expression. In most cases, the gonadal form of CYP19 (CYP19a) was more abundant than the brain form (CYP19b); however, unlike CYP19a, the pattern of CYP19b expression could be clearly segregated into two populations, suggesting an association with sex differentiation. Pharmacological steroids (ethinylestradiol and 17 alpha-methyltestosterone) enhanced the expression of the CYP19b gene at all three days examined (4, 6, and 10 dpf). These data suggest that the timely and appropriate expression of CYP19 is important in development and that the expression of CYP19b (the "extra-gonadal" form) may be associated with sexual differentiation if not sexual determination. J. Exp. Zool. 290:475-483, 2001.

Animals↗
PubMed2001

Hyperlipidemia and reproductive failure in captive-reared alligators: vitamin E, vitamin A, plasma lipids, fatty acids, and steroid hormones.

Blood samples were collected from 26 captive-reared alligators (25 females; one male) and 12 (seven females and five males) wild "nuisance" alligators collected by wildlife personnel in south Louisiana in May 1995. The captive alligators, hatched from artificially incubated eggs in 1972-1973, had received vitamin E supplements during the 3 weeks before the blood sample was collected. Each sample was analyzed for vitamin E (alpha-tocopherol), vitamin A (retinol), total lipid, triacylglycerol, phospholipid, cholesterol, cholesteryl ester, free fatty acids, steroid hormones and a standard clinical blood panel. The fatty acid composition of the plasma lipid fraction was also analyzed. Results indicated that 18 of the captive females and three of the seven wild females were undergoing vitellogenesis, i.e. had elevated plasma estradiol and elevated plasma calcium. Vitellogenic females had higher vitamin E than non-vitellogenic females (77.4 microg/ml vs. 28.6 microg/ml in captive females; 24.0 microg/ml vs. 21 microg/ml in wild females). Plasma retinol was similar in all groups, ranging from 0.5 to 1.4 microg/ml and close to values reported in birds. All lipid fractions, with the exception of cholesteryl ester, were higher in captive alligators than in wild alligators. There were also significant differences in the fatty acid composition of wild and captive alligators. Plasma eicosapentaenoic and docasahexaenoic acid were higher in wild than in captive alligators, whereas linoleic was higher in captive than in wild.

Alligators and Crocodiles↗
PubMed2000

Modulation of arachidonate and docosahexaenoate in Morone chrysops larval tissues and the effect on growth and survival.

The extent to which extreme dietary levels of arachidonate (AA) and/or docosahexaenoate (DHA) modulate lipid composition in the body tissues and consequently affect growth and survival in freshwater Morone larvae species was examined. White bass, M. chrysops, larvae (day 24-46) were fed Artemia nauplii enriched with algal oils containing varying proportions of AA and DHA (from 0 to over 20% the total fatty acids). Growth was significantly reduced (P< 0.05) in larvae fed a DHA-deficient Artemia diet. Increases in dietary levels of AA also were associated with a significant growth reduction. However, the inhibitory effect of AA on larvae growth could be suppressed by the dietary addition of DHA (at a level of 21.6% of the total fatty acids in enrichment lipids). Larval brain + eyes tissue accumulated over 10 times more DHA than AA in its structural lipids (phosphatidylcholine, phosphatidylethanolamine) at any dietary ratio. In contrast, DHA accumulation, as compared to AA, in gill lipids declined considerably at higher than 10:1 DHA/AA tissue ratios. DHA and eicosapentaenoic acid (EPA) contents in brain + eyes tissue were most sensitive to competition from dietary AA, being displaced from the tissue at rates of 0.36 +/- 0.07 mg DHA and 0.46 +/- 0.11 mg EPA per mg increase in tissue AA, and 0.55 +/- 0.14 mg AA per mg increase in tissue DHA. On the other hand, AA and EPA levels in gill tissue were most sensitive to dietary changes in DHA levels; AA was displaced at rates of 0.37 +/- 0.11 mg, whereas EPA increased at rates of 0.68 +/- 0.28 mg per mg increase in tissue DHA. Results suggest that balanced dietary DHA/AA ratios (that allow DHA/AA ratios of 2.5:1 in brain + eyes tissue) promote a high larval growth rate, which also correlates with maximal regulatory response in tissue essential fatty acids.

Animals↗
PubMed1998

Embryonic brain-gonadal axis in temperature-dependent sex determination of reptiles: a role for P450 aromatase (CYP19).

Sex determination in egg-laying amniotes may be fundamentally different from that of placental mammals. The mammalian ovary differentiates normally in the absence of estrogen, whereas estrogen seems to be crucial for proper ovarian development in birds, reptiles, and lower vertebrates. Estrogens are produced normally by the biosynthetic conversion of androgens by the enzyme aromatase (CYP19), which is the sole mediator of this reaction. Aromatase inhibitors are capable of reversing females to males in turtles and chickens; therefore, a role for aromatase as the female sex determinant has been postulated for species in which sex determination is temperature-dependent. The entire aromatase coding sequence (1,509 base pairs) from adult terrapin ovaries was cloned, and Northern analysis indicates a single transcript (2.4 kb) for adult ovaries, whereas male and female brains express a 2.4-kb as well as a 9.6-kb transcript. Using a sensitive (attomole sensitivity) competitive RT-PCR technique, aromatase transcript abundance was quantified during embryonic development for embryos treated with and without estrogen. Aromatase is transcribed, well before the temperature-sensitive, (stage 12), at both male and female temperatures in the brain. There is a switch to lower aromatase transcript abundance in the female brain concurrent with an exponential rise of aromatase transcript in the putative ovary. Transcripts remain below the detection limits in the putative testes but exhibit female levels of aromatase transcript when treated with estrogen. Aromatase mRNA levels are generally reduced in the brain by estradiol application. On the basis of these findings, we have postulated a model based on the competition between 5 alpha-reductase and P450 aromatase for androgen substrate in both the brain and the undifferentiated gonad to explain the TSD phenomenon in reptiles.

Animals↗
PubMed1997

Temperature-dependent aromatase expression in developing diamondback terrapin (Malaclemys terrapin) embryos.

In the diamondback terrapin, Malaclemys terrapin, males hatch at incubation temperatures below 28 degrees C whereas females hatch at temperatures above 30 degrees C. When estrogen is applied to the eggs at male temperatures early in development, females are produced. These data suggest that the enzyme necessary for estrogen synthesis (CYP19, aromatase) in the developing gonad plays a critical role in sex determination in these vertebrates. Accordingly, we have begun an examination of the role and regulation of the aromatase gene in sex determination in the diamond back terrapin, Malaclemys terrapin. We have obtained full-length cDNAs for terrapin ovarian aromatase. Using reverse transcription-polymerase chain reaction (RT-PCR) on mRNA from various tissues we have determined that aromatase is expressed in the female brain and ovary, whereas it is only expressed in the brain of the male. Brain expression of aromatase occurs before stage 15, the beginning of the temperature-dependent sex determining period. Ovarian expression occurs sometime later. To quantify expression levels, we have developed a competitive RT-PCR technique to study the ontogeny of aromatase transcript levels throughout development. The sensitivity of our assay (0.001-10 atmol of transcript) permits us to analyse individual embryonic adrenal/kidney/gonadal complexes without pooling samples. Female hatchlings (stage 26) brains express higher aromatase mRNA levels than male brains (381 +/- 80 vs 202 +/- 85 atmol/microg RNA, respectively). Similarly, ovaries express significantly higher aromatase mRNA levels than hatchling testes (352 +/- 117 vs <0.001 atmol/microg RNA, respectively).

Animals↗
PubMed1994

Role of P-450 aromatase in sex determination of the diamondback terrapin, Malaclemys terrapin.

Sex determination in the diamondback terrapin, Malaclemys terrapin, is temperature-dependent. Eggs incubated at 31 degrees C, and above, hatch in approximately 45 days as females. Eggs incubated below 27 degrees C hatch in about 60 days as males. Sex is not reversible after hatching. Nest temperatures in the wild can be as low as 20 degrees C and as high as 37 degrees C with as much as a 10 degrees C diel cycle. The shortest incubation time measured in nature was 56 days and the longest approaching 120 days. Nests in our study site produced predominantly (> 95%) male hatchlings. Treatment of developing embryos with estrogen produces females at male producing temperatures while treatment with fadrozole (a nonsteroidal aromatase inhibitor) induces partial male-like gonads. Treatment with a steroidal aromatase inhibitor (4-hydroxyandrostenedione, 4-OHA) had no effect on sex determination. Both fadrozole and 4-OHA are potent competitive inhibitors (Ki approximately 40-50 nM) for terrapin in vitro aromatase activity. These findings are consistent with aromatase expression being a key step in sex determination of terrapins. We have cloned a partial single copy P-450 aromatase from the terrapin using a cDNA library constructed from ovarian mRNA. This partial clone is highly homologous to other vertebrate aromatases.

Amino Acid Sequence↗
PubMed1994

Developing intestine is injured during absorption of oleic acid but not its ethyl ester.

Although lipids are essential nutrients in the mammalian diet, we have shown that fatty acids are injurious to epithelial cells of developing piglet intestine during luminal perfusion. Furthermore, the intestine of young animals sustains greater injury than that of older piglets. In an effort to understand the mechanism for this developmental injury, we investigated whether changes in the chemical configuration of oleic acid would alter this damage. Mucosal permeability, as quantitated by the plasma-to-lumen clearance of 51chromium EDTA, was evaluated during luminal perfusion with oleic acid as compared with its ethyl (ethyl oleate) and glyceryl (glycerol-1-mono-oleate) esters, solubilized with taurocholic acid, in jejunum of 1-d-, 3-d-, 2-wk-, and 1-mo-old piglets. 51Chromium EDTA clearance increased significantly during oleic acid and glycerol-1-mono-oleate perfusion, but did not increase during perfusion with ethyl oleate or saline. This result was not secondary to failure of absorption of ethyl oleate, as [14C]oleic acid and ethyl [1-14C]oleate were absorbed to a similar extent. Furthermore, developing intestine was able to remove the ethyl group and then re-esterify the fatty acid to form triacyglycerol. These studies indicate that oleic acid-induced mucosal injury can be abolished when the carboxylic group of the fatty acid is esterified with an ethyl, but not a glycerol, group. Since the ethyl ester is also absorbed and metabolized similarly to the free fatty acid, this may provide a means of supplying long-chain fatty acids to developing intestine without causing mucosal damage.

Animals↗
PubMed1992

Bile is essential for lipid assimilation in Leach's storm petrel, Oceanodroma leucorhoa.

A tritium-labeled glycerol triether was tested as a non-absorbable lipid marker for studies of neutral lipid absorption in normal and cholestryamine-induced steatorrhic chicks of Leach's storm petrel (Oceanodroma leucorhoa). Absorption of triacylglycerol and wax ester (96 +/- 4.2 and 94 +/- 2.1%, respectively) in normal chicks was indistinguishable. Absorption of neutral lipids decreased with increasing quantities of resin, and with the highest dosage (14% wt/wt) less than 30% of dietary lipids were assimilated. The concentration of bile salts (greater than 500 mM) in the gallbladder of Leach's storm petrel is among the highest recorded. The bile salts are predominantly taurine conjugates of chenodeoxycholate and cholate. Biliary cholesterol levels are within the range observed in humans although biliary phospholipid levels are lower and triglyceride levels are higher. Extensive retrograde movement of duodenal contents and biliary secretions to the gizzard and proventriculus were found. Although similarities in fat and bile salt assimilation exist between seabirds and mammals, there do exist differences that make non-polar lipid assimilation (i.e., wax esters) more efficient in seabirds.

Absorption↗
PubMed1992

Comparative aspects of lipid digestion and absorption: physiological correlates of wax ester digestion.

Our understanding of lipid digestion and absorption is largely derived from studies with glyceride-based dietary lipids. Yet for a great majority of marine species, wax esters (long-chain fatty alcohols esterified to long-chain fatty acids) are the dominant dietary neutral lipid. Many birds, especially seabirds (9 species) and some passerines (2 species), have a unique capacity for assimilating wax esters with higher efficiencies (greater than 90%) than that attainable by mammals (less than 50%). This unique capacity is correlated with several factors. One factor is an elevated intestinal bile salt concentration approaching 50 mM and a gallbladder concentration exceeding 600 mM. A second factor involves regular retrograde movement of duodenal contents to the gizzard. Thus not only is gastric emptying closely tied to the receptiveness of the duodenum for further handling of digesta as in mammals, but in birds the reflux returns the digesta (both gastric and duodenal) for further processing to the gizzard. A third key factor in wax ester utilization is a nearly equivalent hydrolysis of wax esters and triglycerides. Although similarities in fat digestion exist between birds and mammals, there do exist differences that make nonpolar lipid assimilation (i.e., wax esters) more efficient in birds, especially for seabirds.

Absorption↗
PubMed1990

Analysis of formaldehyde-induced Adh mutations in Drosophila by RNA structure mapping and direct sequencing of PCR-amplified genomic DNA.

Two formaldehyde-induced mutations at the Drosophila Adh locus (Adhfn45 and Adhfn46) were analyzed by determining RNA structures at different developmental stages, polymerase chain reaction (PCR) amplification of the affected genomic regions, and direct sequencing of the resulting double-stranded DNA fragments. Adhfn46 adults and larvae accumulate abundant ADH-like distal (adult) and proximal (larval) transcripts that are shorter than transcripts in wild-type flies by a lesion located in the second ADH protein-coding exon. Direct sequencing of the amplified DNA region showed that Adhfn46 contains a 69-bp in-frame deletion that removes 23 amino acids near one border of the second exon. Consistent with these findings, we observed a shorter ADHfn46 protein present at only 3% of wild-type levels. In contrast, Adhfn45 adults and larvae accumulate much smaller amounts of ADH-like distal and proximal transcripts. Both RNAs have an identical aberration in RNA splicing of the 65-base intron sequence. Direct sequencing of the amplified mutated DNA region showed that Adhfn45 contains a 21-bp deletion that removed and rearranged DNA at the 5' splice junction of the 65-bp intron. No ADH cross-reacting material is detected in Adhfn45 flies. Direct-repeat sequences (3-11 bp) are present flanking and within the mutated DNA regions. The patterns of DNA deletion and deletion accompanied by sequence addition at the mutant sites suggest a slipped mispairing mechanism during DNA replication or repair that involves local DNA homology.

Alcohol Dehydrogenase↗
PubMed1987

Molecular consequences of two formaldehyde-induced mutations in the alcohol dehydrogenase gene of Drosophila melanogaster.

Adhfn23 and Adhfn24 are two formaldehyde-induced, homozygous-viable, alcohol dehydrogenase-null mutants that bear lesions in the gene that codes for the alcohol dehydrogenase (ADH; EC 1.1.1.1) of Drosophila melanogaster. Adhfn23 contains a 34-base pair deletion in the C-terminal coding region of the alcohol dehydrogenase structural gene. By immunological and molecular analysis, we show that the deletion shifts the translation reading frame and results in a prematurely truncated polypeptide product (10 amino acids shorter than wild type) that cross-reacts with antibody raised against ADH. The steady-state level of alcohol dehydrogenase mRNA present in this mutant is close (97%) to that in the wild type, but the steady-state level of alcohol dehydrogenase-like protein is 50% lower. Moreover, the rate of alcohol dehydrogenase synthesis in Adhfn23 flies is reduced to 60% of that found in the wild type. Hence both the rate of synthesis and the rate of degradation of alcohol dehydrogenase are affected. In contrast, Adhfn24 which contains an 11-base pair deletion in the N-terminal coding region of the ADH gene, synthesizes no immunodetectable protein, and the amount of alcohol dehydrogenase mRNA is less than half that of wild-type flies. As with Adhfn23, the deletion in Adhfn24 results in a change in the reading frame. Unlike Adhfn23, however, nucleic acid sequence data indicate that polypeptide chain elongation can proceed for a considerable distance (over 130 amino acids) beyond the deletion. Based upon antigenic binding-site predictions, the resultant aberrant protein (projected 195 amino acids in length) would share few antigenic sites with the alcohol dehydrogenase from the wild type, which may account for the lack of immunoprecipitable material in this mutant. The contrasting effects these two deletions have on the Drosophila ADH mRNA levels and ADH protein levels are discussed.

Alcohol Dehydrogenase↗
PubMed1987

Reexamination of alcohol dehydrogenase structural mutants in Drosophila using protein blotting.

Using protein blotting and an immuno-overlay procedure, we have reexamined the cross-reacting material produced by ADH null-activity mutants generated with ethyl methanesulfonate (EMS). Of the 13 mutants, 11 have an immunodetectable polypeptide of wild-type size. The native and urea denatured isoelectric points (pI) establish that 7 of 13 of the mutations have no effect on protein charge. The electrophoretic mobilities of each variant on increasing percent acrylamide gels (Ferguson analysis), reveal that 9 of the 11 immunodetectable mutants have retained the ability to form dimers under native conditions. None of the inactive mutant proteins has the ability to form the "adduct-bound" isozyme. We have found no correlation between protein pI and in vivo stability. The observed frequencies of specific charge class alterations do not dispute the propensity of G:A transitions previously found for EMS mutagenesis.

Alcohol Dehydrogenase↗
PubMed1987

Partial correction of structural defects in alcohol dehydrogenase through interallelic complementation in Drosophila melanogaster.

Alcohol dehydrogenases (ADH) from the F1 progeny of all pairwise crosses between 12 null-activity mutants and crosses between these mutants and four active variants, ADHn5 ADHF, ADHD and ADHS, were analyzed for the presence of active or inactive heterodimers. Gels were stained for ADH enzyme activity, and protein blots of duplicate gels were probed with ADH-specific antibody to detect cross-reacting material. Crosses between the three major electrophoretic variants. ADHF, ADHS and ADHD, all produced active heterodimers. Four mutant proteins (ADHn2, ADHn4, ADHn10 and ADHn13) did not form heterodimers with any other ADH subunit tested. Of the 28 crosses involving the remaining null activity mutants, 22 produce heterodimers. Twelve of these exhibit partial restoration of enzyme activity. In five cases of active heterodimers from null-activity crosses, Adhn11 supplied one of the subunits. In two crosses involving the active variant ADHD, the null activity mutant subunits (ADHn8 and ADHn3) destabilized the heterodimer sufficiently to cause inactivation of the ADHD subunit. In the cross between AdhF and Adhn3, the activity of the ADHF subunit was also greatly reduced in association with the ADHn3 subunit. Two crosses (Adhn1 X Adhn11 and Adhn5 X Adhn12) result in partial restoration of one of the homodimeric proteins (ADHn1 and ADHn12, respectively), as well as forming active heterodimers.

Alcohol Dehydrogenase↗