HIV-1 LTR C/EBP binding site sequence configurations preferentially encountered in brain lead to enhanced C/EBP factor binding and increased LTR-specific activity.

Recent studies have shown that two CAAT/enhancer binding protein (C/EBP) sites are critically important for efficient human immunodeficiency virus (HIV) type 1 (HIV-1) replication within cells of the monocyte/macrophage lineage, a primary cell type infected by HIV-1 and a potentially important vehicle for transport of virus to the central nervous system (CNS). Given the relevance of HIV-1 LTR sequence variation with respect to HIV-1 replication within monocyte populations and the important role that monocyte tropism likely plays in HIV-1 infection of the brain, C/EBP site sequence variation was examined within peripheral blood- and brain-derived LTR populations. Brain-derived LTRs commonly possessed a C/EBP site I configuration (6G, comprised of a thymidine to guanosine substitution with respect to the clade B consensus sequence at position 6 of C/EBP site I) that leads to enhanced binding of C/EBP proteins over that observed with the HIV-1 clade B consensus sequence at this site. In contrast, the 6GC/EBP site I configuration appeared infrequently within sequenced peripheral blood-derived LTRs. In addition, C/EBP site II was even more highly conserved in brain-derived HIV-1 LTR populations than site I. This was not the case with peripheral blood-derived LTR C/EBP site II sequences. The high degree of C/EBP site II conservation in brain-derived LTRs was likely important in LTR regulation since the clade B consensus sequence conserved at C/EBP site II recruited high amounts of C/EBP family members. Transient transfection analyses indicated that conservation of the strong C/EBP site II in brain-derived LTRs was likely due to important interactions with Tat. Overall, brain-derived HIV-1 LTRs preferentially contained two highly reactive C/EBP binding sites, which may suggest that these sites play important roles in LTR-directed transcription during invasion and maintenance of HIV-1 in the central nervous system.

Interaction between CCAAT/enhancer binding protein and cyclic AMP response element binding protein 1 regulates human immunodeficiency virus type 1 transcription in cells of the monocyte/macrophage lineage.

Recent observations have shown two CCAAT/enhancer binding protein (C/EBP) binding sites to be critically important for efficient human immunodeficiency virus type 1 (HIV-1) replication within cells of the monocyte/macrophage lineage, a cell type likely involved in transport of the virus to the brain. Additionally, sequence variation at C/EBP site I, which lies immediately upstream of the distal nuclear factor kappa B site and immediately downstream of a binding site for activating transcription factor (ATF)/cyclic AMP response element binding protein (CREB), has been shown to affect HIV-1 long terminal repeat (LTR) activity. Given that C/EBP proteins have been shown to interact with many other transcription factors including members of the ATF/CREB family, we proceeded to determine whether an adjacent ATF/CREB binding site could affect C/EBP protein binding to C/EBP site I. Electrophoretic mobility shift analyses indicated that selected ATF/CREB site variants assisted in the recruitment of C/EBP proteins to an adjacent, naturally occurring, low-affinity C/EBP site. This biophysical interaction appears to occur via at least two mechanisms. First, low amounts of CREB-1 and C/EBP appear to heterodimerize and bind to a site consisting of a half site from both the ATF/CREB and C/EBP binding sites. In addition, CREB-1 homodimers bind to the ATF/CREB site and recruit C/EBP dimers to their cognate weak binding sites. This interaction is reciprocal, since C/EBP dimer binding to a strong C/EBP site leads to enhanced CREB-1 recruitment to ATF/CREB sites that are weakly bound by CREB. Sequence variation at both C/EBP and ATF/CREB sites affects the molecular interactions involved in mediating both of these mechanisms. Most importantly, sequence variation at the ATF/CREB binding site affected basal LTR activity as well as LTR function following interleukin-6 stimulation, a treatment that leads to increases in C/EBP activation. Thus, HIV-1 LTR ATF/CREB binding site sequence variation may modulate cellular signaling at the viral promoter through the C/EBP pathway.

Analysis of the HIV-1 LTR NF-kappaB-proximal Sp site III: evidence for cell type-specific gene regulation and viral replication.

It has been widely demonstrated that the human immunodeficiency virus type 1 (HIV-1) envelope, specifically the V3 loop of the gp120 spike, evolves to facilitate adaptation to different cellular populations within an infected host. Less energy has been directed at determining whether the viral promoter, designated the long terminal repeat (LTR), also exhibits this adaptive quality. Because of the unique nature of the cell populations infected during the course of HIV-1 infection, one might expect the opportunity for such adaptation to exist. This would permit select viral species to take advantage of the different array of conditions and factors influencing transcription within a given cell type. To investigate this hypothesis, the function of natural variants of the NF-kappaB-proximal Sp element (Sp site III) was examined in human cell line models of the two major cell types infected during the natural course of HIV-1 infection, T cells and monocytes. Utilizing the HIV-1 LAI molecular clone, which naturally contains a high-affinity Sp site III, substitution of low-affinity Sp sites in place of the natural site III element markedly decreased viral replication in Jurkat T cells. However, these substitutions had relatively small effects on viral replication in U-937 monocytic cells. Transient transfections of HIV-1 LAI-based LTR-luciferase constructs into these cell lines suggest that the large reduction in viral replication in Jurkat T cells, caused by low-affinity Sp site III variants, may result from reduced basal as well as Vpr- and Tat-activated LTR activities in Jurkat T cells compared to those in U-937 monocytic cells. When the function of Sp site III was examined in the context of HIV-1 YU-2-based LTR-luciferase constructs, substitution of a high-affinity element in place of the natural low-affinity element resulted in increased basal YU-2 LTR activity in Jurkat T cells and reduced activity in U-937 monocytic cells. These observations suggest that recruitment of Sp family members to Sp site III is of greater importance to the function of the viral promoter in the Jurkat T cell line as compared to the U-937 monocytic cell line. These observations also suggest that other regions of the LTR may compensate for Sp recruitment defects in specific cell populations.

Effect of single benzo[a]pyrene diol epoxide-deoxyguanosine adducts on the action of DNA polymerases in vitro.

Neither Sequenase 2.0 nor Klenow fragment were able to extend 12-mer primers using the eight templates (16-mers) derived by placing each of the four isomeric benzo[a]pyrene diol epoxide-deoxyguanosine adducts at the 13th nucleotide from the 3'-end of two different sequence contexts. Using an 11-mer primer to get a running start did not overcome the adduct induced block of primer extension except for the Klenow fragment and one of the two sequence contexts, indicating primer extension is dependent on both the polymerase and sequence context. In this case, purine nucleoside triphosphates (dATP>dGTP) were incorporated opposite each of the four adducts.

Effects of administrative license revocation on employment.

This study addresses the impact of administrative license revocation (ALR) on the employment and income of first and multiple drunk-driving (DUI) offenders. It also inquires into the impact of alcohol-related crashes on the employment of other persons involved in such crashes (i.e. innocent drivers, passengers and pedestrians). Questionnaires were completed by 579 first-time offenders and 233 multiple offenders at alcohol education schools and treatment programs in four counties in four states, which represented varying ALR laws: Chester County, PA (without ALR); Anne Arundel County, MD (ALR with immediate hardship license available); Marin County, CA (30-day hard license suspension); and New Castle County, DE (90-day hard license suspension). Completed crash 'victim' surveys were returned by 146 crash victims from Pennsylvania, California and Delaware. The study found that ALR does not have a major impact on the DUI offender's job and income. Alcohol-involved crashes can have a great impact on seriously injured victims, but the proportion of DUI crashes producing serious injury is quite low. Most DUI is crash-free, and most crashes do not involve injury. The vast bulk of the impact of DUI events falls on the offenders rather than innocent victims.

Detection of chromosomal aneuploidy in endometriosis by multi-color fluorescence in situ hybridization (FISH).

Endometriosis affects 10-15% of women of reproductive age and is a common cause of infertility and pelvic pain. Although endometriosis is characterized by abnormal growth or turn-over of cells, the genetic changes involved remain unclear. We employed a multi-color fluorescence in situ hybridization (FISH) strategy to determine the incidence of somatic chromosomal numeric alterations in severe/late stage endometriosis. Using alpha-satellite sequence-specific DNA probes for chromosomes 7, 8, 11, 12, 16, 17, and 18, simultaneous two- and three-color FISH were performed to evaluate the frequency of monosomic, disomic, and trisomic cells in normal control and endometriotic tissue specimens. In one of four endometriosis samples studied, a significantly higher frequency of monosomy for chromosome 17 (14.8%, chi 2(4) = 53.3, P < 0.0001) and 16 (8.8%, chi 2(4) = 11.4, P < 0.05) was observed. An increased number of cells with chromosome 11 trisomy (14.8%, chi 2(4) = 96.2, P < 0.0001) were detected in a second case. In a third case, a distinct colony of nuclei with chromosome 16 monosomy (14.1%, chi 2(4) = 21.39, P < 0.005) was detected. Acquired chromosome-specific aneuploidy may be involved in endometriosis, reflecting clonal expansion of chromosomally abnormal cells. That candidate tumor suppressor genes and oncogenes have been mapped to chromosomes 11, 16, and 17 suggests that chromosomal loss or gain plays a role in the development and/or progression of endometriosis.

Severe clinical phenotype due to an interstitial deletion of the short arm of chromosome 1: a brief review.

We report on a newborn girl with malformed ears, bilateral cleft lip and cleft palate, complex congenital heart disease, absent left thumb, and rib abnormalities. Cytogenetic analysis demonstrated a de novo interstitial deletion of the short arm of chromosome 1 [46,XX,del(1)(p21p22.3)]. Reports of interstitial deletions on the short arm of chromosome 1 are rare. However, when comparing this patient's phenotype to others with deletions of 1p, we found that the current case was much more severely affected than previously reported cases.

Maternal serum screening for fetal genetic disorders.

Maternal serum analyte screening is an integral component of contemporary antenatal care. Based on elevated MSAFP levels, 85% to 90% of NTDs (e.g., anencephaly or spina bifida) can be detected. Using a combination of serum analytes (e.g., MSAFP, hCG, UE3) 55% to 60% of fetal Down's syndrome can be detected. Future strategies for Down's syndrome screening may include the use of new markers such as dimeric inhibin-A and urinary beta-core fragment of hCG, as well as first-trimester screening, particularly with PAPP-A and free beta-hCG.

Mutagenic specificities and adduct distributions for 7-bromomethylbenz[a]anthracenes.

Mutation induction in the supF gene of the plasmid pS189 by 7-bromomethylbenz[a]anthracene and 7-bromomethyl-12-methylbenz[a]anthracene was examined. The former compound was substantially more mutagenic than the latter but a much greater proportion of the total mutations were located at mutation hotspots for the 12-methyl derivative. The overall correlation between sites of mutation and sites of polymerase arrest (an indicator of adduct formation) through the supF gene was poor. Although these bromocompounds should form only a single guanine adduct (unlike dihydrodiol epoxides that form both cis and trans adducts) more than one mutational change was found at a given site, although the predominant base substitution was G-->T for either compound.

License plate confiscation for persistent alcohol impaired drivers.

Minnesota cancels the registrations and confiscates the license plates of vehicles driven by repeat drinking drivers in a procedure which prior research has demonstrated to be effective in reducing subsequent recidivism. The research reported here concerns the problems experienced in the functioning of this law. Samples of officials and repeat driving under the influence (DUI) offenders were interviewed in Minnesota and in the neighboring state of Iowa, chosen for comparison because its laws also provide for plate confiscation, but using a judicial rather than an administrative procedure. In addition, representative samples of the driving and vehicle registration records of convicted drunk drivers and of routine traffic offenders were analyzed. The research found that evasion of plate impoundment orders by drivers, though apparently easy to accomplish, appeared to be rare. However, the orders were themselves not issued in a large proportion of cases where they were prescribed by statute, potentially weakening the effectiveness of the law. The reasons, with possible countermeasures, are explored in this report.

Abandonment of mandatory jail for impaired drivers in Norway and Sweden.

In 1988 and 1990, respectively, Norway and Sweden adopted legal reforms including abandonment of mandatory jail sentences for persons driving with BACs above specific limits. Interrupted time-series analysis finds that in both countries traffic deaths diminished simultaneously with the reforms, consistent with the understanding that Scandinavian success in reducing impaired driving does not depend upon mandatory jail.

Reaction with DNA and mutagenic specificity of syn-benzo[g]chrysene 11,12-dihydrodiol 13,14-epoxide.

The spectroscopic characterization of purine deoxyribonucleoside adducts derived from the fjord-region syn-benzo[g]chrysene 11,12-dihydrodiol 13,14-epoxide and the mutagenic specificity of the latter compound for the supF gene in the pSP189 shuttle vector are described. This dihydrodiol epoxide preferentially forms adducts with deoxyadenosine residues in DNA and is preferentially opened trans in reactions with DNA or with deoxyribonucleotides. In common with other fjord-region syn-dihydrodiol epoxides, the most frequently observed mutational changes were A-->T and G-->T changes. This hydrocarbon dihydrodiol epoxide is structurally similar to syn-benzo[c]phenanthrene 3,4-dihydrodiol 1,2-epoxide but has an additional benzene ring annelated distant from the reaction center. As anticipated, there were some common features in the chemistry and mutagenicities of these two compounds, but there were also substantive differences which indicate factors of importance in controlling reactions of these kinds of compounds with DNA.

Comparison of the sites of reaction of three polycyclic aromatic hydrocarbons in the supF gene.

The distribution of hydrocarbon-DNA adducts through the supF gene in plasmid pS189 was examined using the polymerase arrest assay. For three hydrocarbon dihydrodiol epoxides, derived from 5-methylchrysene, 7-methylbenz[a]anthracene, and benzo[a]pyrene, that exhibit a preference for reaction with guanine residues in DNA, polymerase arrest spectra were similar but not identical. For each agent, guanines in different sequence contexts exhibited varying reactivities and each specific guanine did not necessarily respond to each agent in the same fashion. Thus, sequence context together with the individual dihydrodiol epoxide's chemical and physical properties all play a role in determining sites and extents of reaction within a specific gene. The polymerase arrest data were not predictive of the known sites of mutation hotspots for these dihydrodiol epoxides in the supF gene indicating that further action upon the adducted DNA by repair systems is probably necessary to determine which specific chemical adducts will ultimately give rise to mutation.

Prevalence of alcohol-impaired driving: an international comparison.

This paper reports international differences in the extent of alcohol-impaired driving. These differences are then interpreted in the light of inducements and disincentives to drink and to drive.

Punishment as a factor in preventing alcohol-related accidents.

As a humanitarian and economic 'bad', infliction of punishment requires justification in terms of compensating achievements, as well as moral appropriateness. In the context of road accidents, there is evidence that increasing the certainty and swiftness of threatened punishment may deter risky behavior, although increasing the severity of the threat seems ineffective. These effects may be generalizable to other kinds of accidents, but empirical evidence is lacking. A further possibility for the justifiable use of punishment is in its application to negligent service of alcohol.

Drunk driving among American blacks and Hispanics.

This paper reports the results of a literature search to determine the extent and nature of minority group involvement in drunk driving. Most of the research supports the view that American blacks and Hispanics are disproportionately more likely to be drunk drivers. However, the evidence is not fully consistent. The general relationship seems to be reduced or even reversed for minority youth. Furthermore, studies based on self-reported behavior contradict those based on official statistics such as alcohol-related deaths, traffic arrests, and accidents. Self reports generally show less drunk driving among the minority groups.

Effects of license revocation on drunk-driving offenders.

Revoking or suspending the licenses of persons convicted of driving while intoxicated has often been claimed to be uniquely effective in reducing subsequent dangerous driving, as indexed by crashes and violations. This finding holds despite the impression that many suspended and revoked drivers continue to drive illegally. The research described here inquires into the operation of license actions. It uses intensive personal interviews with offenders to obtain information concerning the nature and amount of illegal driving and the effects of license deprivation on employment and other aspects of the offenders' lives.