Use of suppressive subtractive hybridization and cDNA arrays to discover patterns of altered gene expression in the liver of dihydrotestosterone and 11-ketotestosterone exposed adult male largemouth bass (Micropterus salmoides)

In this study male largemouth bass (LMB) were exposed to the naturally occurring androgens, dihydrotestosterone (DHT) or 11-ketotestosterone (11-KT) in order to identify genes that are differentially regulated by these steroid hormones. Using subtractive hybridization on livers of fish treated with DHT against vehicle control, many novel LMB genes were cloned. These genes were added to our gene library and arrayed. Six genes were up-regulated and five were down-regulated by both androgens. But, each androgen also regulated specific genes. One gene that was identified as a potential androgen marker was spermidine-spermine-N(1)-acetyltransferase that was up-regulated by both androgens. Determining which genes are responsive to natural androgens will help to identify biochemical pathways that are impacted.     

Induction of zona radiata and vitellogenin genes in estradiol and nonylphenol exposed male sheepshead minnows (Cyprinodon variegatus)

Several genes normally induced by estradiol (E(2)) in female fish, those for vitellogenins (VTGs) and zona radiata proteins (ZRPs), are also inducible in males exposed to estrogenic chemicals. Male sheepshead minnows (SHM) were exposed to both E(2) and para-nonylphenol (NP), at several doses and times to determine a dose-response. Quantitative real time PCR was used to measure mRNA for VTG1, VTG2, ZRP2 and ZRP3. Both E(2) and NP elicited a dose-response increase in all of the mRNAs tested. Exposure to both chemicals resulted in VTG2 expression at about a 10-fold lower level than VTG1, and ZRP2 expression at a lower level than ZRP3.     

Array technology as a tool to monitor exposure of fish to xenoestrogens

A variety of anthropogenic chemicals are capable of binding to the estrogen receptor of vertebrate species. Binding of these compounds can interfere with homeostasis by disrupting normal gene expression patterns. The purpose of this study was to investigate the feasibility of applying array technology as a monitoring tool for detecting the presence and distribution of estrogenic compounds in coastal habitats using sheepshead minnows as our model. cDNA clones that were isolated from differential display, including vitellogenin alpha and beta, vitelline envelope protein (ZP2), and transferrin, among others, were spotted on the macroarray. The results of these experiments demonstrate a characteristic expression pattern of estrogen responsive genes in sheepshead minnows exposed to 17 beta-estradiol (E2).     

Use of suppressive subtractive hybridization and cDNA arrays to discover patterns of altered gene expression in the liver of dihydrotestosterone and 11-ketotestosterone exposed adult male largemouth bass (Micropterus salmoides)

In this study male largemouth bass (LMB) were exposed to the naturally occurring androgens, dihydrotestosterone (DHT) or 11-ketotestosterone (11-KT) in order to identify genes that are differentially regulated by these steroid hormones. Using subtractive hybridization on livers of fish treated with DHT against vehicle control, many novel LMB genes were cloned. These genes were added to our gene library and arrayed. Six genes were up-regulated and five were down-regulated by both androgens. But, each androgen also regulated specific genes. One gene that was identified as a potential androgen marker was spermidine-spermine-N¹-acetyltransferase that was up-regulated by both androgens. Determining which genes are responsive to natural androgens will help to identify biochemical pathways that are impacted.     

Impacts of cyclic hypoxia on reproductive and gene expression patterns in the grass shrimp: field versus laboratory comparison

Identification of organisms able to act as biological indicators of hypoxia exposure is important given the increasing frequency of hypoxic events worldwide. Grass shrimp, Palaemonetes pugio, are ubiquitous estuarine residents in the southeastern US, where they commonly experience cyclic hypoxia. We hypothesized that reproductive and gene expression patterns of grass shrimp in cyclic hypoxic field sites relative to normoxic sites would be similar to previous results from cyclic hypoxia laboratory experiments showing differential up- and down-regulation of hypoxia-responsive genes. There were few differences in gene expression of indigenous shrimp collected during summer from two Gulf of Mexico estuarine systems [East Bay, FL (EB) and Weeks Bay, AL (WB)], although down-regulation of the gene coding for vitellogenin-1 corresponded with a decreased relative fecundity and fewer ovigerous females at cyclic hypoxic field sites, suggesting an overall impact on reproduction. The gene expression profiles of grass shrimp caged for 7 days in field sites differed by estuary, but few hypoxia-responsive genes identified in laboratory studies were differentially expressed in field shrimp. Overall, genes coding for protein synthesis, protein degradation, carbohydrate and lipid metabolism and electron transport were mostly up-regulated in EB caged shrimp but generally down-regulated in WB caged shrimp and laboratory shrimp. Thus, caged grass shrimp from different bay systems exhibited profoundly different gene expression profiles. Such profiles may serve as sensitive bioindicators of differences in water quality, habitat quality or food resources among estuaries but are not effective as indicators of hypoxia exposure.     

Induction of zona radiata and vitellogenin genes in estradiol and nonylphenol exposed male sheepshead minnows (Cyprinodon variegatus)

Several genes normally induced by estradiol (E₂) in female fish, those for vitellogenins (VTGs) and zona radiata proteins (ZRPs), are also inducible in males exposed to estrogenic chemicals. Male sheepshead minnows (SHM) were exposed to both E₂ and para-nonylphenol (NP), at several doses and times to determine a dose-response. Quantitative real time PCR was used to measure mRNA for VTG1, VTG2, ZRP2 and ZRP3. Both E₂ and NP elicited a dose-response increase in all of the mRNAs tested. Exposure to both chemicals resulted in VTG2 expression at about a 10-fold lower level than VTG1, and ZRP2 expression at a lower level than ZRP3.