Transcriptional activity of the genes for apoproteins A-I and E in neonatal rat liver.

The expression of the genes for apoproteins A-I and E (apoA-I, apoE) in fetal and neonatal rat livers was quantitated by measurement of steady-state levels of mRNA and by assays of relative transcriptional activity using the nuclear 'run-on' technique. From the last day of gestation (day -1) to day 5 after birth (day 5), for apoE there was a 2.2-fold increase in relative transcriptional activity and a 2.6-fold increase in steady-state mRNA. For apoA-I, from day -1 to 3 there was a 1.25-fold increase in both transcription and steady-state mRNA levels. We conclude that the increase in steady-state mRNA for the apoE gene which occurs during liver development in the rat is facilitated primarily by transcriptional control.

Anticoccidial derivatives of 6-azauracil. 5. Potentiation by benzophenone side chains.

Attachment of p-benzophenone side chains at N1 was found to be one of the most effective modifications for enhancing the potency of 6-azauracil against a broad spectrum of coccidia in chickens. Compound 20 was about 1000-fold more potent than 6-azauracil. Structure-activity relationships paralleled those found in a previously reported series of related analogues containing diphenyl sulfide and sulfone side chains. Drug metabolism studies showed the ketones to be reduced rapidly to carbinols, which are the prevalent species in vivo.

A novel family of cell surface receptors with tyrosine kinase-like domain.

Human cDNA clones encoding two novel proteins with a region strongly homologous to the tyrosine kinase domain of growth factor receptors, in particular of the Trk family, were obtained by a polymerase chain reaction-based approach. These proteins, Ror1 and Ror2, share 58% overall amino acid identity and a structure indicative of cell surface molecules. A secretion signal sequence and a transmembrane domain delimit the extracellular portion, which contains immunoglobulin-like, cysteine-rich, and kringle domains. The cytoplasmic portion contains the tyrosine kinase-like domain which (in Ror2) appears to be associated with protein kinase activity in vitro, followed by serine/threonine- and proline-rich motifs. Partial nucleotide sequences of the rat genes reveal striking evolutionary conservation of the proteins between human and rat. The level of expression of the rat genes is high in the head and body of early embryo and decreases dramatically after embryonic day 16. Based on these data, Ror1 and Ror2 appear to define a new developmentally regulated family of cell surface receptors for unidentified ligands.

In vitro mutagenesis of the herpes simplex virus type 1 DNA polymerase gene results in altered drug sensitivity of the enzyme.

A mutation (asparagine 815 to serine 815) was introduced into the herpes simplex virus type 1 (HSV-1) DNA polymerase (pol). The HSV-1 pol enzyme in lysates of Saccharomyces cerevisiae cells expressing the mutant protein showed increased resistance to acyclovir triphosphate and increased sensitivity to phosphonoacetate but was not substantially altered with respect to sensitivity to phosphonoformate or aphidicolin. These results directly demonstrate that both resistance to acyclovir triphosphate and sensitivity to phosphonoacetate can be conferred by this mutation in the absence of other viral factors and that the yeast expression system can be used for structure-function studies on HSV-1 pol.

Structure-function studies of the herpes simplex virus type 1 DNA polymerase.

The analysis of the deduced amino acid sequence of the herpes simplex virus type 1 (HSV-1) DNA polymerase reported here suggests that the polymerase structure consists of domains carrying separate biological functions. The HSV-1 enzyme is known to possess 5'-3'-exonuclease (RNase H), 3'-5'-exonuclease, and DNA polymerase catalytic activities. Sequence analysis suggests an arrangement of these activities into distinct domains resembling the organization of Escherichia coli polymerase I. In order to more precisely define the structure and C-terminal limits of a putative catalytic domain responsible for the DNA polymerization activity of the HSV-1 enzyme, we have undertaken in vitro mutagenesis and computer modeling studies of the HSV-1 DNA polymerase gene. Sequence analysis predicts that the major DNA polymerization domain of the HSV-1 enzyme will be contained between residues 690 and 1100, and we present a three-dimensional model of this region, on the basis of the X-ray crystallographic structure of the E. coli polymerase I. Consistent with these structural and modeling studies, deletion analysis by in vitro mutagenesis of the HSV-1 DNA polymerase gene expressed in Saccharomyces cerevisiae has confirmed that certain amino acids from the C terminus (residues 1073 to 1144 and 1177 to 1235) can be deleted without destroying HSV-1 DNA polymerase catalytic activity and that the extreme N-terminal 227 residues are also not required for this activity.