PubMed Health⌕ Search

Biomedical subjects

S L Levine

Publications and source records attributed to S L Levine.

14 recordsLinked to original sources
PubMed2002

Okadaic acid increases ARNT homodimer transactivation potential.

The human aryl hydrocarbon nuclear translocator (hARNT) protein belongs to the family of basic helix-loop-helix (bHLH) PAS transcription factors and regulates a range of cellular processes by either homodimerizing or heterodimerizing with other bHLH-PAS proteins. hARNT has been shown to be almost exclusively phosphorylated on serine residues. However, regulation of hARNT with respect to phosphorylation remains poorly understood. The phosphatase inhibitor okadaic acid was used to explore whether a change in hARNT phosphorylation status could influence hARNT homodimer activity. The hARNT homodimer has been shown to bind to E boxes and E-box binding factors are believed to be important in the regulation of cell differentiation and proliferation. Okadaic acid significantly increased hARNT-mediated class B, E-box-driven reporter activity in COS-1 cells, transiently expressing hARNT without affecting hARNT protein levels. This alteration in hARNT-mediated class B, E-box-driven reporter activity correlates with an observed increase in [32P]orthophosphate incorporation into hARNT. Treatment with okadaic acid resulted in a 12-fold increase in [32P]orthophosphate incorporation into hARNT that was transiently expressed in COS-1 cells; an increase in the number of tryptic phosphopeptides generated from hARNT digests on two-dimensional phosphopeptide maps was also observed. Despite the significant increase in [32P]orthophosphate incorporation into hARNT, serine remained the predominantly phosphorylated residue. Clearly, increased serine phosphorylation does not appear to negatively regulate hARNT homodimerization or transactivation potential. These results demonstrate that increased hARNT homodimer signaling in COS-1 cells may result from a direct change in hARNT phosphorylation status.

Animals↗
PubMed2001

Aryl hydrocarbon receptor (AhR)/AhR nuclear translocator (ARNT) activity is unaltered by phosphorylation of a periodicity/ARNT/single-minded (PAS)-region serine residue.

The aryl hydrocarbon nuclear translocator (ARNT) protein belongs to the family of basic helix-loop-helix (HLH)-periodicity/ARNT/single-minded [Per/ARNT/Sim (PAS)] transcription factors and regulates a range of cellular processes by either homodimerizing or heterodimerizing with other basic HLH-PAS proteins. To date, it has been shown that both the HLH and PAS domains are required for aryl hydrocarbon receptor (AhR) ARNT heterodimerization and that phosphorylation of ARNT is also required for this heterodimerization. Presently, regulation of ARNT with respect to phosphorylation is poorly understood. In an earlier study, murine ARNT was shown to be a phosphoprotein, to display charge heterogeneity, and to have a shift in its predominant isoforms after heterodimerization with the AhR. It was hypothesized that this shift may represent a change in ARNT phosphorylation status. Metabolic [(32)P]orthophosphate labeling of human ARNT-transfected COS-1 cells, in conjunction with phosphoamino acid analysis, Edman degradation, and phosphopeptide mapping, demonstrated that ARNT is predominantly phosphorylated on serine residues and that serine 348 (S348) in the PAS domain is phosphorylated. Alanine and glutamic acid substitutions were used to demonstrate that loss of phosphorylation at this site did not influence AhR-mediated xenobiotic response elements-driven or ARNT-mediated class B E-box-driven signaling. Additionally, the phosphorylation pattern of ARNT was unaltered after AhR heterodimerization. Although phosphorylation of S348 did not modulate AhR-ARNT or ARNT-ARNT signaling, phosphorylation of this PAS-region serine residue may be important in other ARNT-mediated gene expression systems.

Amino Acid Sequence↗
PubMed2000

A tetratricopeptide repeat half-site in the aryl hydrocarbon receptor is important for DNA binding and trans-activation potential.

Similar to certain unliganded steroid hormone receptor complexes, the unliganded aryl hydrocarbon receptor has been shown to consist of a multimeric core complex that includes the 90-kDa heat shock protein (hsp90) and the immunophilin-like hepatitis B X-associated protein 2 (XAP2). Immunophilins and XAP2 associated with these complexes bind to the carboxyl-terminal end of hsp90 through an interaction with their tetratricopeptide repeat (TPR) domains. The consensus TPR binding motif contains two domains, A and B. Recently, the carboxyl terminus of XAP2 has been shown to contain a highly conserved TPR domain that is required for the assembly of XAP2 with both hsp90 and AhR. A search of the murine AhR sequence identified domain B (A-F-A-P) of the consensus TPR sequence directly adjacent to the carboxyl-terminal side of the helix-loop-helix region of the murine and human AhR. We hypothesized that this conserved domain B region may be involved with mediating interactions between either AhR-hsp90, AhR-XAP2, and/or AhR-AhR nuclear translocator protein. Site-directed mutagenesis of the amino-terminal alanine residue of this region to an aspartic acid (A78D) completely inhibited 2,3,7, 8-tetrachloro-p-dioxin (TCDD) -dependent activation of a xenobiotic response element (XRE) driven gene expression construct in transfected COS-1 and BP8 cells. The A82F mutation caused a 40 to 50% decrease in TCDD-dependent activation. The inability of A78D and the reduction of A82F to trans-activate XRE-driven reporter activity did not result from impaired AhR-XAP2-hsp90 interactions, TCDD-dependent AhR translocation to the nucleus, or AhR-AhR nuclear translocator protein interactions. In vitro DNA binding analysis demonstrated that loss of trans-activation potential by the A78D mutation resulted from impaired XRE binding. This study underscores the potential importance of AhR mutations that occur naturally outside of known functional domains.

Amino Acid Sequence↗
PubMed1999

Noncompetitive mixed-type inhibition of rainbow trout CYP1A catalytic activity by clotrimazole.

Over the past two decades a number of antifungal imidazole derivatives have been approved for use in agricultural. The purpose of this study was to characterize the interaction of a model antifungal imidazole compound with a cytochrome P450 isozyme in a species of fish. Clotrimazole inhibited rainbow trout (Oncorhyncus mykiss) hepatic CYP1A-catalyzed ethoxyresorufin O-deethylase (EROD) activity in vivo and in vitro. Although clotrimazole inhibited EROD activity in vivo, it did not effect CYP1A mRNA levels. Addition of clotrimazole to microsomes produced a type II binding spectrum and clotrimazole was determined to be a noncompetitive mixed-type inhibitor of EROD activity with an IC50 of 190 nM. Since antifungal imidazole compounds may be co-applied with other pesticides, inhibition of cytochrome P450 activity by antifungal imidazole compounds may lead to unexpected toxicological interactions.

Animals↗
PubMed1997

Induction of CYP1A mRNA and catalytic activity in gizzard shad (Dorosoma cepedianum) after waterborne exposure to benzo[a]pyrene.

The purpose of this study was to examine the concentration and time-dependent response of hepatic CYP1A mRNA levels and ethoxyresorufin O-deethylase (EROD) activity in gizzard shad (Dorosoma cepedianum) after waterborne exposure to the polycyclic aromatic hydrocarbon benzo[a]pyrene (BaP). Gizzard shad hepatic CYP1A mRNA levels and EROD activity were increased within 6 hr by waterborne concentrations of 1.01 microgram BaP/l and were highest after 72 hr of exposure. The median effective time to reach 50% of maximum induction (ET50) values for CYP1A mRNA and EROD activity was 12 +/- 9 and 34 +/- 3 hr, respectively. The rise of CYP1A mRNA levels ahead of the rise of EROD activity showed that the CYP1A gene has the general pattern of a gene that is under transcriptional control. Although CYP1A mRNA levels were maximally induced after 72 hr of exposure, they returned to basal levels between 72 and 120 hr of exposure. Despite a decrease in CYP1A mRNA levels between 72 and 120 hr of exposure, EROD activity did not significantly change between 72 and 120 hr of exposure. These results indicate that hepatic CYP1A mRNA levels in gizzard shad can only feasibly identify short-term exposures to BaP concentrations of approximately 1 microgram/l and that are not longer than 72 hr. Consequently, hepatic CYP1A catalytic activity (e.g., EROD) and CYP1A protein levels should be measured in addition to hepatic CYP1A mRNA levels when biomonitoring aquatic systems for certain types of chemical contamination.

Animals↗
PubMed1996

Relations of motor and vocal milestones in typically developing infants and infants with Down syndrome.

We measured the ages at which typically developing infants and infants with Down syndrome achieved an important vocal milestone (canonical babbling), a rhythmic motor milestone (hand-banging), and six other motor milestones. The interrelations of the milestone onsets, and their relations with Down syndrome, were assessed quantitatively. Hand-banging and canonical babbling were associated and were somewhat delayed by Down syndrome. Stepping, standing, sitting, and creeping/crawling were associated and were severely delayed by Down syndrome. Rolling and reaching were also delayed by Down syndrome, though they were not strongly associated with other milestones or with one another. These results suggest that the rhythmic behaviors (canonical babbling and hand-banging) may be internally linked by common neuromuscular underpinnings and that the postural behaviors may be similarly linked.

Child Development↗
PubMed1995

Structure/function studies of mammalian Na-H exchangers--an update.

Four mammalian Na+/H+ exchangers have recently been cloned. Despite the structural similarity, these Na+/H+ exchanger isoforms differ in kinetic characteristics and their response to external stimuli. The present review deals with the recent developments in their functional characterization and their short-term regulation.

Animals↗
PubMed1995

Onset of speech-like vocalizations in infants with Down syndrome.

Canonical babbling of infants with and without Down syndrome was compared. Infants with Down syndrome and typically developing infants began canonical babbling in the first year of life, but the infants with Down syndrome began 2 months later. Once begun, their canonical babbling was less stable than that of typically developing infants. Age at onset of canonical babbling for the infants with Down syndrome was correlated with their scores at 27 months of age on the Early Social-Communication Scales. The results of this study suggest that Down syndrome influences vocal development in the first year of life and that early vocal development is related, possibly in combination with motoric and cognitive factors, to later social and communicative functioning of children with Down syndrome.

Child, Preschool↗