Phillip A. Sharp

Bio: Phillip A. Sharp is an academic researcher from Massachusetts Institute of Technology. The author has contributed to research in topics: RNA & RNA splicing. The author has an hindex of 172, co-authored 614 publications receiving 117126 citations. Previous affiliations of Phillip A. Sharp include McGovern Institute for Brain Research & Medical Research Council.

RNA splicing and genes.

Abstract: The splicing of long transcripts of RNA (copied from DNA in the cell nucleus) into smaller, specific mRNA (ready for export to the protein-producing machinery in the cytoplasm) is an important event in the regulation of gene expression in eukaryotic cells. The splicing reaction occurs as a late step in the nuclear pathway for synthesis of mRNAs. This pathway commences with initiation of transcription by RNA polymerase II and probably involves an integrated series of steps each dependent on previous events. Splicing of precursors to mRNAs involves the formation of a spliceosome complex containing the 5' and 3' splice sites. This complex contains the evolutionarily highly conserved small nuclear RNAs (snRNAs) U2, U4, U5, and U6. The most abundant snRNA, U1, is required to form the spliceosome and may be a part of the spliceosome. Analogues of these snRNAs have been identified in yeast. Assembly of the spliceosome probably involves the binding of a multi-snRNA complex containing U4, U5, and U6 snRNAs. Several observations suggest that the association of snRNAs in such complexes is quite dynamic. It is argued that the snRNAs in the spliceosome form a catalytic RNA structure that is responsible for the cleavage and ligation steps during splicing. ( JAMA 1988;260:3035-3041)

Adenovirus transcription. IV. Synthesis of viral-specific RNA in human cells infected with temperature-sensitive mutants of adenovirus 5.

Abstract: Cytoplasmic RNA sequences produced in HeLa cells infected with the adeno-virus 5 temperature-sensitive mutants ts1, ts2, ts9, ts17, ts18, ts19, ts20, ts22, ts49, ts36, and ts125 were characterized by hybridization to DNA probes generated by strand separation of restriction endonuclease fragments of adenovirus 5 DNA. Two ""early'' mutants defective in DNA synthesis, ts125 and ts36, fail to make wild-type levels of all previously reported classes of late RNA at the nonpermissive temperature. At 40.5 degrees C, both ts125 and ts36 synthesize a wild-type complement of early cytoplasmic RNA 16 h after infection. Under these conditions, no ""late'' cytoplasmic RNA sequences were observed. Similarly, nuclear RNA present in these cells resembled early cytoplasmic RNA rather than late nuclear RNA. All the late adenovirus 5 temperature-sensitive mutants synthesized normal wild-type levels of late cytoplasmic RNA at the nonpermissive temperature, except ts2, which appears to overproduce certain cytoplasmic species.

Studies on the molecular nature of r factors

Abstract: There are at least five potential experimental approaches to the problem of control of transferable drug resistance in pathogenic bacteria: (1) a search for new antimicrobials; (2) elimination of R factors by selective inhibition of R-factor replication; (3) interference with genetic expression of R-factors, either at the level of gene transcription or at the stage of translation of messenger RNA into protein; (4) prevention of formation of new R factors; and ( 5 ) inhibition of transfer of drug resistance. Although the first approach can be largely empirical, rational use of the last four approaches depends upon a fundamental understanding of extrachromosomal drug resistance at a molecular level. With this in mind, we have been attempting to elucidate the molecular nature of antibiotic resistance factors in the Enterobacteriaceae. Considerable controversy has existed about the molecular nature of R factors.'D2 Transduction experiments carried out in Salmonella and E. coli almost a decade ago led Watanabe and FukasawaS to propose that R factors consist of a transfer unit (RTF segment) that controls transmission of the plasmid, linearly linked, as shown in FIGURE la, to drug resistance determinants which carry genetic information specifying resistance to antimicrobial agents. More recent studies by Anderson and his collaborators,\" and later by Mitsuhashi and coworkers,5 have shown that the transfer units of at least certain classes of R factors can be transmitted alone as well as in combination with resistance determinants. Furthermore, these investigators have found that ordinarily nontransmissible R determinants can interact with transfer units (although not necessarily covalently as depicted in this Figure), which are then able to mediate their passage by conjugation (FIGURE la) . Once within a recipient cell, the linked transfer and resistance units can replicate autonomously. The earliest studies of the molecular nature of R factorsas7 indicated that heterogeneous satellite bands of DNA are associated with the presence of R factors in Proteus mirabilis and certain other bacterial species. However, the methods of nucleic acid extraction used in these early experiments yielded DNA having a maximum molecular weight of 10-15 million daltons, whereas the molecular weight of R-factor DNA in virro was estimated at more than three times this size.6 Because the R factor was not intact in these studies, it could not be determined whether the several R-factor satellite bands identified by cesium chloride gradient centrifugation reflected intramolecular heterogeneity within a single fragmented R-factor species, as in the case of bacteriophage X,8 or whether there were in fact functionally associated, but physically distinct, R-factor subunits. More recently, procedures have been developed that have made it possible to obtain high molecular weight DNA from bacteria by detergent lysis and phenol

Methods and options for estimating iron and zinc bioavailability using Caco-2 cell models: Benefits and limitations

Abstract: Micronutrient deficiencies are prevalent worldwide and have detrimental effects on human health. Complex interactions between micronutrients and other dietary components largely determine micronutrient bioavailability, and understanding these interactions is key to improving micronutrient status. A number of in vitro and in vivo methodologies are available for assessing micronutrient bioavailability. The purpose of this review is to highlight the usefulness of one of the in vitro models, the Caco-2 cell, as a predictive tool for human micronutrient bioavailability. The review focuses on current methods used with the Caco-2 cell line, their benefits and limitations, and the possibilities for the future development of this model.

Initial sequencing and analysis of the human genome.

Abstract: The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Abstract: 抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

A rapid alkaline extraction procedure for screening recombinant plasmid DNA

Abstract: A procedure for extracting plasmid DNA from bacterial cells is described. The method is simple enough to permit the analysis by gel electrophoresis of 100 or more clones per day yet yields plasmid DNA which is pure enough to be digestible by restriction enzymes. The principle of the method is selective alkaline denaturation of high molecular weight chromosomal DNA while covalently closed circular DNA remains double-stranded. Adequate pH control is accomplished without using a pH meter. Upon neutralization, chromosomal DNA renatures to form an insoluble clot, leaving plasmid DNA in the supernatant. Large and small plasmid DNAs have been extracted by this method.