A superfamily of S locus-related sequences in Arabidopsis: diverse structures and expression patterns.

Six sequences that are closely related to the S gene family of the largely self-incompatible Brassica species have been identified in self-fertilizing Arabidopsis. The sequences define four genomic regions that map to chromosomes 1 and 3. Of the four functional genes identified, only the previously reported Arabidopsis AtS1 gene was expressed specifically in papillar cells and may function in pollination. The remaining three genes, including two novel genes designated ARK2 and ARK3, encode putative receptor-like serine/threonine protein kinases that are expressed predominantly in vegetative tissues. ARK2 promoter activity was detected exclusively in above-ground tissues, specifically in cotyledons, leaves, and sepals, in correlation with the maturation of these structures. ARK3 promoter activity was detected in roots as well as above-ground tissues but was limited to small groups of cells in the root-hypocotyl transition zone and at the base of lateral roots, axillary buds, and pedicels. The nonoverlapping patterns of expression of the ARK genes and the divergence of their sequences, particularly in their predicted extracellular domains, suggest that these genes perform nonredundant functions in specific aspects of development or growth of the plant body.

Distinct cis-acting elements direct pistil-specific and pollen-specific activity of the Brassica S locus glycoprotein gene promoter.

The promoter of the S Locus Glycoprotein (SLG) gene of Brassica is a tightly regulated promoter that is active specifically in reproductive organs. In transgenic tobacco, this promoter is active exclusively in cells of the pistil and in pollen. We transformed tobacco with truncated versions of the SLG13 promoter fused to the beta-glucuronidase reporter gene. We show that the promoter has a modular organization and consists of separable DNA elements that independently specify pistil- and pollen-specific expression. A 196-bp region (-339 to -143) is sufficient to confer stigma and style specificity to the marker gene. Two distinct, but functionally redundant, domains (-415 to -291 and -117 to -8) allow specific expression of the gene in pollen. The functional domains identified within the SLG13 promoter contain sequence elements that are highly conserved in different alleles of the SLG gene and in the S Locus Related SLR1 gene.

Structure and expression of AtS1, an Arabidopsis thaliana gene homologous to the S-locus related genes of Brassica.

Genetic and molecular analysis of the self-incompatibility locus (S-locus) of the crucifer Brassica has led to the characterization of a multigene family involved in pollen-stigma interactions. While the crucifer Arabidopsis thaliana does not have a self-incompatibility system, S-related sequence were detected in this species by cross-hybridization with Brassica DNA probes. In this paper, we show that an A. thaliana S-related sequence, designated AtS1, is expressed specifically in flower buds. Sequence analysis suggests that AtS1 encodes a secreted glycoprotein that is most similar to the Brassica S-locus related protein SLR1. As has been proposed for SLR1, this gene may be involved in determining some fundamental aspect of pollen-stigma interactions during pollination. The molecular and genetic advantages of the Arabidopsis system will provide many avenues for testing this hypothesis.

Brassica S-Proteins Accumulate in the Intercellular Matrix along the Path of Pollen Tubes in Transgenic Tobacco Pistils.

A tobacco plant transformed with a Brassica oleracea SLG-22 gene was analyzed by immunocytochemical methods to determine the localization of the transgene-encoded protein product. Immunolabeling was observed in the pistil along the path followed by pollen tubes after pollination. S-antigen accumulated in the intercellular matrix of the transmitting tissue of the style and its continuation in the basal portion of the stigma and outside a few special cells of the placental epidermis of the ovary. This pattern of S-antigen distribution closely resembles that described for the S-associated glycoproteins of self-incompatible Nicotiana alata and differs from its distribution in B. oleracea.

A highly conserved Brassica gene with homology to the S-locus-specific glycoprotein structural gene.

The S-locus-specific glycoprotein of Brassica and the gene encoding it (the SLG gene) are thought to be involved in determining self-incompatibility phenotype in this genus. It has been shown that the Brassica genome contains multiple SLG-related sequences. We report here the cloning and characterization of a Brassica oleracea gene, SLR1, which corresponds to one of these SLG-related sequences. Like the SLG gene, SLR1 is developmentally regulated. It is maximally expressed in the papillar cells of the stigma at the same stage of flower development as the onset of the incompatibility response. Unlike SLG, the SLR1 genes isolated from different S-allele homozygotes are highly conserved, and this gene, which appears to be ubiquitous in crucifers, is expressed in self-compatible strains as well as self-incompatible strains. Most importantly, we show that the SLR1 gene is not linked to the S-locus and therefore cannot be a determinant of S-allele specificity in Brassica.

Mapping Pieris rapae granulosis virus transcripts and their in vitro translation products.

Poly(A)+ RNA was isolated from Pieris rapae granulosis virus (PrGV)-infected P. rapae larvae at times early (40 h postinoculation) and late (88 h postinoculation) in larval infection. Using Northern (RNA) blot analysis, we determined the sizes, relative abundances, and map locations of over 100 PrGV transcripts. Differences were found in the transcripts which had accumulated at the two time points. Splicing of these transcripts was not detected. Evidence for the expression of overlapping RNAs in PrGV was obtained. A minimum of 35 PrGV translation products were detected via hybrid selection of poly(A)+ RNA with specific PrGV restriction endonuclease fragments, followed by in vitro translation.

Replication of the Trichoplusia ni granulosis and nuclear polyhedrosis viruses in cell cultures.

Newly established Trichoplusia ni (cabbage looper) embryonic cell lines were infected with T. ni granulosis virus (TnGV) and T. ni nuclear polyhedrosis virus (TnSNPV). Infection of cultured cells with TnGV was ascertained by peroxidase-antiperoxidase staining, DNA slot-blot hybridization, and transmission electron microscopy. Initially, 15 cell lines supported TnGV replication, the percentage of infected cells ranging from 1 to 50%.However, susceptibility of the 15 cell lines to TnGV infection either decreased or was lost within 20 to 25 passages from the initial primary culture. Infection of cells with TnSNPV was determined by phase contrast and electron microscopy. TnSNPV infected 29 36 cell lines tested, the percentage of infected cells ranging from 1 to 60%.