Ryotaro Ishizaki

Immunological properties of avian oncornavirus polypeptides

Abstract The major polypeptides and glycoproteins of avian myeloblastosis virus and of the Prague strain of Rous sarcoma virus were isolated by gel filtration in guanidine hydrochloride (GuHCl). Hyperimmune sera prepared in rabbits against homogeneous preparations of each material were used to study the nature of the antigenic specificities on these molecules. The results indicated that (1) each component contained unique antigenic determinants; (2) each of four polypeptides (27,000, 19,000, 15,000, 12,000 daltons) contained group-specific (gs) reactivity; and (3) subgroup specific reactivity was found in the 19,000-dalton polypeptide.

Normal chicken cells (chf−) express a surface antigen which cross-reacts with determinants of the major envelope glycoprotein (gp85) of avian myeloblastosis virus☆

Abstract Using rabbit antisera raised against the purified major envelope glycoprotein (gp85) of avian myeloblastosis virus (AMV), we were unable to differentiate between normal chicken cells carrying the chicken-helper factor (chf+) from those lacking this endogenous virus information (chf−) in several serological assays, including competition radioimmunoassay and membrane immunofluorescence. Radioimmunoprecipitation analysis of the material recognized in uninfected chf(+) and chf(-) chicken cells by the rabbit anti-AMV gp85 antisera revealed the presence in immune precipitates of both cell types of a high molecular weight glycoprotein (∼125,000), while low amounts of gp85 were precipitated only from the chf(+) cells. In contrast, antisera raised against the gp85s of three other avian oncornaviruses did not recognize the 125,000-dalton molecule in either cell type, although gp85 was again precipitated from the chf(+) chicken cells. The high molecular weight glycoprotein (denoted NCG) appears to represent a normal chicken cell surface glycoprotein which is antigenically cross-reactive with unique determinants of AMV gp85, presumably dependent on the carbohydrate portion of the molecules. The significance of this cross-reactivity is discussed, as well as the interpretation of previous results obtained through the use of these rabbit anti-AMV gp85 antisera.

The Structural Polypeptides of Equine Infectious Anemia Virus

The structural polypeptides and glycoproteins of equine infectious anemia virus were identified following electrophoresis in SDS-PAGE. The major non-glycosylated polypeptides had molecular weights of 25,000, 14,000 and 11,000 daltons. Two glycoproteins of 80,000 and 40,000 daltons were also detected. The relationship of these components to the structural elements of mammalian C-type oncoviruses is discussed.

Restriction Fragment Length Polymorphisms Detected in N-ras-Related Sequences of Rats and Their Linkage Analyses

Novel restriction fragment length polymorphisms (RFLPs) in inbred rats were revealed with the human N-ras gene as probe. Three fragments hybridizing to the probe were detected by Southern blot hybridization under highly stringent conditions, and one of the fragments showed variation in inbred rat strains. Furthermore, on hybridization under low-stringency conditions, an additional fragment hybridizing to the probe was observed, and this fragment also showed interstrain variation. These two variant fragments showed different distributions in 27 inbred rat strains and segregated in backcross progeny as codominant alleles of independent single autosomal loci. Therefore, the loci for these RFLPs were namedNras-1 andNras-2, respectively. Analyses of linkages between the RFLPs and 11 other loci revealed that theNras-2 locus was closely linked to thec locus (3.7±2.6%), which belongs to rat linkage group I.

Novel restriction fragment length polymorphism of the growth hormone gene in inbred rats

A novel restriction fragment length polymorphism in inbred rats was detected by Southern blot analysis with rat growth hormone cDNA as a probe. Four alleles, characterized byPstI fragments of 1.2, 1.1, 0.9, and 0.7 kb, respectively, were detected in 27 strains examined. The same distribution of polymorphisms was observed on digestion of DNAs of these strains with three other enzymes,PvuII,HindIII, andBamHI. Moreover, the same differences in length of allelic restriction fragments were obtained with these restriction enzymes as withPstI. These findings suggested that the polymorphism was caused by insertion or deletion of variable DNA segments in the second intron of the growth hormone gene. Linkage analyses using backcross progeny provided no evidence for close linkage between the restriction fragment length polymorphism locus and 10 other loci examined.

Restriction Fragment Length Polymorphism of Cardiac Myosin Heavy Chain Gene in Rats and Its Strain Distribution

The distribution of an RFLP in EcoRI fragments of the cardiac myosin heavy chain gene among 29 strains of laboratory rats was examined. Southern blot hybridization of rat genomic DNAs with rat cardiac myosin heavy chain cDNA as a probe demonstrated an interstrain variation in one of eight EcoRI fragments. Of the 28 inbred strains examined, 10 had a fragment of 10 kbp, whereas 18 had a fragment of 7.5 kbp. The 15 samples of the remaining strain (Iar: WI outbred stock) had fragments of either 7.5 kbp or 10 and 7.5 kbp, indicating that this strain has maintained heterogeneity of these fragments.