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Biomedical subjects

N Hernandez

Publications and source records attributed to N Hernandez.

At least 19 recordsLinked to original sources
PubMed2003

Predictability of electron cone ratios with respect to linac make and model.

In the past, the Radiological Physics Center (RPC) has developed standard sets of photon depth-dose and wedge-factor data, specific to the make, model, and wedge design of the linear accelerator (linac). In this paper, the RPC extends the same concept to electron-cone ratios. Since 1987, the RPC has measured and documented cone-ratio (CR) values during on-site dosimetry review visits to institutions participating in National Cancer Institute cooperative clinical trials. Data have been collected for approximately 500 electron beams from a wide spectrum of linac models. The analysis presented in this paper indicates that CR values are predictable to 2% to 3% (two standard deviations) for a given make and model of linac with a few exceptions. The analysis also revealed some other interesting systematics. For some models, such as the Varian Clinac 2500 and the Elekta/Philips SL18, SL20, and SL25, CR values were nearly identical for cone sizes 15 cm x 15 cm (or 14 cm x 14 cm) and 20 cm x 20 cm across the range of available energies. Certain models of the same make of linac, such as the Mevatron MD, KD, and 6700 series models or the Clinac 2100 and 2300 models, exhibited indistinguishable CRs. Irrespective of linac model, two consistent general trends were observed: namely, an increase in CR value with incident beam energy for cone sizes smaller than 10 cm x 10 cm and a decrease with energy for cone sizes larger than 10 cm x 10 cm. These data are valuable to the RPC as a quality assurance remote-monitoring tool to identify potential dosimetry errors. The physics community will also find the data useful in several ways: as a redundant check for clinical values in use, to validate the values measured during commissioning of new machines or to ensure consistency of values measured during annual quality assurance procedures.

Electrons↗
PubMed2001

Reconstitution of transcription from the human U6 small nuclear RNA promoter with eight recombinant polypeptides and a partially purified RNA polymerase III complex.

The human U6 small nuclear (sn) RNA core promoter consists of a proximal sequence element, which recruits the multisubunit factor SNAP(c), and a TATA box, which recruits the TATA box-binding protein, TBP. In addition to SNAP(c) and TBP, transcription from the human U6 promoter requires two well defined factors. The first is hB", a human homologue of the B" subunit of yeast TFIIIB generally required for transcription of RNA polymerase III genes, and the second is hBRFU, one of two human homologues of the yeast TFIIIB subunit BRF specifically required for transcription of U6-type RNA polymerase III promoters. Here, we have partially purified and characterized a RNA polymerase III complex that can direct transcription from the human U6 promoter when combined with recombinant SNAP(c), recombinant TBP, recombinant hB", and recombinant hBRFU. These results open the way to reconstitution of U6 transcription from entirely defined components.

Amino Acid Sequence↗
PubMed2001

A positioned nucleosome on the human U6 promoter allows recruitment of SNAPc by the Oct-1 POU domain.

The human snRNA promoters contain a proximal sequence element (PSE) required for basal transcription and a distal sequence element (DSE) required for activated transcription. The PSE recruits the multisubunit factor SNAPc, whereas the DSE recruits Oct-1. Oct-1 and SNAPc bind cooperatively to DNA when their respective binding sites are moved into proximity through a mechanism that involves a defined protein-protein contact. Here, we show that on the natural U6 promoter, cooperative binding of Oct-1 and SNAPc is mediated by a positioned nucleosome that resides between the DSE and the PSE. This cooperative binding requires the same protein-protein contact as cooperative binding to closely spaced sites on naked DNA and mediates transcription activation.

Allosteric Site↗
PubMed2001

Relationships of sexual, physical, and emotional abuse to emotional and behavioral problems among incarcerated adolescents.

This study examined the relationships of sexual, physical and emotional abuse to emotional (internalizing) and behavioral (externalizing) problems among incarcerated girls and boys. Participants were youth who were remanded to the correctional facilities within a statewide juvenile correctional system in a southern state in the United States of America. Each participant completed a structured interview regarding abuse history, emotional and behavioral difficulties, and demographic characteristics. Multiple regression analyses indicated that girls were more likely than boys to internalize their problems. The only abuse variable that was positively and significantly associated with emotional problems was emotional abuse. Greater behavioral problems were significantly related to youths being younger in age, white ethnicity, history of sexual abuse, and history of physical abuse. There were overall gender differences for internalizing problems, but not for externalizing problems among incarcerated adolescents. Furthermore, physical and sexual abuses were related to externalizing problems but not to internalizing problems. Thus, different types of abuse appear to have different effects on adolescent behavior. Implications for future research and practice are discussed.

Adolescent↗
PubMed2000

A map of protein-protein contacts within the small nuclear RNA-activating protein complex SNAPc.

The nucleation of RNA polymerases I-III transcription complexes is usually directed by distinct multisubunit factors. In the case of the human RNA polymerase II and III small nuclear RNA (snRNA) genes, whose core promoters consist of a proximal sequence element (PSE) and a PSE combined with a TATA box, respectively, the same multisubunit complex is involved in the establishment of RNA polymerase II and III initiation complexes. This factor, the snRNA-activating protein complex or SNAP(c), binds to the PSE of both types of promoters and contains five types of subunits, SNAP190, SNAP50, SNAP45, SNAP43, and SNAP19. SNAP(c) binds cooperatively with both Oct-1, an activator of snRNA promoters, and in the RNA polymerase III snRNA promoters, with TATA-binding protein, which binds to the TATA box located downstream of the PSE. Here we have defined subunit domains required for SNAP(c) subunit-subunit association, and we show that complexes containing little more than the domains mapped here as required for subunit-subunit contacts bind specifically to the PSE. These data provide a detailed map of the subunit-subunit interactions within a multifunctional basal transcription complex.

Amino Acid Sequence↗
PubMed2000

Different human TFIIIB activities direct RNA polymerase III transcription from TATA-containing and TATA-less promoters.

Transcription initiation at RNA polymerase III promoters requires transcription factor IIIB (TFIIIB), an activity that binds to RNA polymerase III promoters, generally through protein-protein contacts with DNA binding factors, and directly recruits RNA polymerase III. Saccharomyces cerevisiae TFIIIB is a complex of three subunits, TBP, the TFIIB-related factor BRF, and the more loosely associated polypeptide beta("). Although human homologs for two of the TFIIIB subunits, the TATA box-binding protein TBP and the TFIIB-related factor BRF, have been characterized, a human homolog of yeast B(") has not been described. Moreover, human BRF, unlike yeast BRF, is not universally required for RNA polymerase III transcription. In particular, it is not involved in transcription from the small nuclear RNA (snRNA)-type, TATA-containing, RNA polymerase III promoters. Here, we characterize two novel activities, a human homolog of yeast B("), which is required for transcription of both TATA-less and snRNA-type RNA polymerase III promoters, and a factor equally related to human BRF and TFIIB, designated BRFU, which is specifically required for transcription of snRNA-type RNA polymerase III promoters. Together, these results contribute to the definition of the basal RNA polymerase III transcription machinery and show that two types of TFIIIB activities, with specificities for different classes of RNA polymerase III promoters, have evolved in human cells.

Amino Acid Sequence↗
PubMed2000

Variations in prudent laypersons' perceptions of the need for emergent medical care.

The objective of this study was to determine whether differences exist among distinct demographic groups' tendencies to view various medical symptoms as emergent in nature. A questionnaire containing six demographic questions and listing 29 common medical complaints was distributed to 306 respondents who agreed to participate; they were asked to indicate when (if at all) they would be likely to seek medical care for each problem. Overall, across the series of complaints, African American respondents desired statistically significantly more acute care than Caucasians. Those with a high school education or lower also wanted to be seen statistically significantly sooner for more symptoms than those with some college education or higher. We conclude that opinions as to the urgency of medical symptoms vary widely among demographic groups.

Adult↗
PubMed1999

SNAP(c): a core promoter factor with a built-in DNA-binding damper that is deactivated by the Oct-1 POU domain.

snRNA gene transcription is activated in part by recruitment of SNAP(c) to the core promoter through protein-protein contacts with the POU domain of the enhancer-binding factor Oct-1. We show that a mini-SNAP(c) consisting of a subset of SNAP(c) subunits is capable of directing both RNA polymerase II (Pol II) and Pol III snRNA gene transcription. Mini-SNAP(c) cannot be recruited by Oct-1, but binds as efficiently to the promoter as SNAP(c) together with Oct-1 and directs activated RNA Pol III transcription. Thus, SNAP(c) represses its own binding to DNA, and repression is relieved by interactions with the Oct-1 POU domain that promote cooperative binding. We have shown previously that TBP also represses its own binding, and in that case repression is relieved by cooperative interactions with SNAP(c). This may represent a general mechanism to ensure that core promoter-binding factors, which have strikingly slow off-rates, are recruited specifically to promoter sequences rather than to cryptic-binding sites in the genome.

DNA-Binding Proteins↗
PubMed1999

FBI-1, a factor that binds to the HIV-1 inducer of short transcripts (IST), is a POZ domain protein.

The HIV-1 promoter directs the synthesis of two classes of transcripts, short, non-polyadenylated transcripts and full-length, polyadenylated transcripts. The synthesis of short transcripts is activated by a bipartite DNA element, the inducer of short transcripts or IST, located downstream of the HIV-1 transcriptional start site, while the synthesis of full-length transcripts is activated by the viral activator Tat. Tat binds to the RNA element TAR, which is encoded largely between the two IST half-elements. Upon activation by Tat, the synthesis of short RNAs is repressed. We have previously purified a factor called FBI-1 (for factor that binds to IST) whose binding to wild-type and mutated ISTs correlated well with the abilities of these ISTs to direct the synthesis of short transcripts. Here, we report the cloning of cDNAs encoding FBI-1. FBI-1 contains a POZ domain at its N-terminus and four Krüppel-type zinc fingers at its C-terminus. The C-terminus is sufficient for specific binding, and FBI-1 can form homomers through its POZ domain and, in vivo, through its zinc finger domain as well. In addition, FBI-1 associates with Tat, suggesting that repression of the short transcripts by Tat may be mediated through interactions between the two factors.

Amino Acid Sequence↗
PubMed1999

The general transcription factors IIA, IIB, IIF, and IIE are required for RNA polymerase II transcription from the human U1 small nuclear RNA promoter.

RNA polymerase II transcribes the mRNA-encoding genes and the majority of the small nuclear RNA (snRNA) genes. The formation of a minimal functional transcription initiation complex on a TATA-box-containing mRNA promoter has been well characterized and involves the ordered assembly of a number of general transcription factors (GTFs), all of which have been either cloned or purified to near homogeneity. In the human RNA polymerase II snRNA promoters, a single element, the proximal sequence element (PSE), is sufficient to direct basal levels of transcription in vitro. The PSE is recognized by the basal transcription complex SNAPc. SNAPc, which is not required for transcription from mRNA-type RNA polymerase II promoters such as the adenovirus type 2 major late (Ad2ML) promoter, is thought to recruit TATA binding protein (TBP) and nucleate the assembly of the snRNA transcription initiation complex, but little is known about which GTFs other than TBP are required. Here we show that the GTFs IIA, IIB, IIF, and IIE are required for efficient RNA polymerase II transcription from snRNA promoters. Thus, although the factors that recognize the core elements of RNA polymerase II mRNA and snRNA-type promoters differ, they mediate the recruitment of many common GTFs.

Animals↗
PubMed1998

The Oct-1 POU domain activates snRNA gene transcription by contacting a region in the SNAPc largest subunit that bears sequence similarities to the Oct-1 coactivator OBF-1.

The RNA polymerases II and III snRNA gene promoters contain an octamer sequence as part of the enhancer and a proximal sequence element (PSE) as part of the core promoter. The octamer and the PSE bind the POU domain activator Oct-1 and the basal transcription factor SNAPc, respectively. Oct-1, but not Oct-1 with a single E7R mutation within the POU domain, binds cooperatively with SNAPc and, in effect, recruits SNAPc to the PSE. Here, we show that SNAPc recruitment is mediated by an interaction between the Oct-1 POU domain and a small region of the largest subunit of SNAPc, SNAP190. This SNAP190 region is strikingly similar to a region in the B-cell-specific Oct-1 coactivator, OBF-1, that is required for interaction with octamer-bound Oct-1 POU domain. The Oct-1 POU domain-SNAP190 interaction is a direct protein-protein contact as determined by the isolation of a switched specificity SNAP190 mutant that interacts with Oct-1 POU E7R but not with wild-type Oct-1 POU. We also show that this direct protein-protein contact results in activation of transcription. Thus, we have identified an activation target of a human activator, Oct-1, within its cognate basal transcription complex.

Amino Acid Sequence↗
PubMed1998

SNAP19 mediates the assembly of a functional core promoter complex (SNAPc) shared by RNA polymerases II and III.

The basal transcription factor SNAPc binds to the PSE, a core element in the RNA polymerase II and III human snRNA promoters. SNAPc contains at least four subunits, but it has not been possible to assemble a fully defined recombinant SNAPc. Here we reconstitute SNAPc from five recombinant subunits, SNAP43, SNAP45, SNAP50, SNAP190, and a newly identified subunit, SNAP19. This recombinant complex binds specifically to the PSE and directs both RNA polymerase II and III snRNA gene transcription. Thus, the same core SNAPc nucleates the assembly of two classes of initiation complexes.

Amino Acid Sequence↗
PubMed1998

The HIV-1 inducer of short transcripts activates the synthesis of 5,6-dichloro-1-beta-D-benzimidazole-resistant short transcripts in vitro.

The HIV-1 inducer of short transcripts (IST) is an unusual promoter element that activates the synthesis of short transcripts from the HIV-1 promoter as well as from heterologous promoters. While the DNA sequences constituting IST have been characterized in some detail, little is known about the biochemical mechanisms underlying IST activity. Here, we describe a cell-free transcription assay that faithfully reproduces the synthesis of IST-dependent HIV-1 short transcripts. As in vivo, formation of these short transcripts requires a functional IST element and is repressed in the presence of the viral trans-activator Tat. Short transcript and full-length transcript synthesis respond differently to variations in several reaction parameters, suggesting that the short and full-length transcripts are synthesized by transcription complexes with distinct biochemical properties. In particular, short transcript synthesis is resistant to the action of 5,6-dichloro-1-beta-D-benzimidazole, an inhibitor of transcript elongation. Formation of transcription complexes directed by the IST element may, therefore, not require the activity of a factor inhibited by 5, 6-dichloro-1-beta-D-benzimidazole, such as the TFIIH-associated or pTEFb kinases.

Base Sequence↗
PubMed1998

The large subunit of basal transcription factor SNAPc is a Myb domain protein that interacts with Oct-1.

The human RNA polymerase II and III snRNA promoters have similar enhancers, the distal sequence elements (DSEs), and similar basal promoter elements, the proximal sequence elements (PSEs). The DSE, which contains an octamer motif, binds broadly expressed activator Oct-1. The PSE binds a multiprotein complex referred to as SNAPc or PTF. On DNAs containing both an octamer site and a PSE, Oct-1 and SNAPc bind cooperatively. SNAPc consists of at least four stably associated subunits, SNAP43, SNAP45, SNAP50, and SNAP190. None of the three small subunits, which have all been cloned, can bind to the PSE on their own. Here we report the isolation of cDNAs corresponding to the largest subunit of SNAPc, SNAP190. SNAP190 contains an unusual Myb DNA binding domain consisting of four complete repeats (Ra to Rd) and a half repeat (Rh). A truncated protein consisting of the last two SNAP190 Myb repeats, Rc and Rd, can bind to the PSE, suggesting that the SNAP190 Myb domain contributes to recognition of the PSE by the SNAP complex. SNAP190 is required for snRNA gene transcription by both RNA polymerases II and III and interacts with SNAP45. In addition, SNAP190 interacts with Oct-1. Together, these results suggest that the largest subunit of the SNAP complex is involved in direct recognition of the PSE and is a target for the Oct-1 activator. They also provide an example of a basal transcription factor containing a Myb DNA binding domain.

Amino Acid Sequence↗
PubMed1998

Evaluation of in vivo models for studying calcification behavior of commercially available bovine pericardium.

BACKGROUND AND AIM OF THE STUDY: A common frame of reference is essential when attempting to determine if new treatments intended to reduce calcification of bioprostheses are superior to existing processes and products. The aim of this study was to examine calcification behavior for a commercially available pericardial bioprosthesis in subcutaneous and sheep valve models, and to evaluate the importance of appropriate control treatments in comparative studies with proposed new treatments. METHODS: Samples of bovine pericardium were placed subcutaneously under the dorsal skin of weanling rats and juvenile rabbits for 30-, 60- and 90-day intervals. Samples were either commercially available pericardial tissue or tissue processed with phosphate-buffered glutaraldehyde alone. Commercially available pericardial valves were also implanted in the mitral position in juvenile sheep, with elective sacrifice at 20 weeks. Retrieved samples underwent X-ray, histologic and elemental analysis. RESULTS: Commercial samples retrieved from the subcutaneous and sheep models showed similar, minimal calcification behavior on X-ray and histologic slides, whereas pericardium exposed to glutaraldehyde alone demonstrated rapid calcification. CONCLUSIONS: The 90-day subcutaneous rabbit model produced patterns of calcification similar to those in valves explanted from juvenile sheep after 20 weeks. A statistically significant decrease (p < 10(-8)) in calcification was demonstrated for clinical pericardium when compared with pericardium exposed to glutaraldehyde alone in the subcutaneous model. This suggests that subcutaneous models may be a cost-effective, time-efficient means of evaluating and comparing various tissue treatment methods. The rabbit methodology may provide a more accurate prediction of clinical performance, offering a greater degree of sensitivity. These studies also indicate that the commercially available process shows minimal calcification in the commonly used 30-day weanling rat subcutaneous model, contradicting other reported studies that may not accurately represent commercially available processes.

Animals↗