Developmental expression of genetically defined peptidases in maize

The activities of genetically defined amino- and endopeptidases of maize were compared in pericarp, endosperm, and embryonic tissue of the maize kernel from 5 days postpollination until harvest. Activities were highest in the immature stages and declined as drying progressed. The expression of some of the peptidase genes contributed by the pollen parent was examined during early endosperm development in an F₁ cross. The paternally contributed peptidase variants could first be detected 7 days after pollination.     

Gene activation of alcohol dehydrogenase in danio hybrids

The regulation of alleles encoding the enzyme alcohol dehydrogenase (ADH) was investigated in F1Brachydanio hybrids (zebra danio female x spotted danio male) by acrylamide gel electrophoresis. Both parental species showed a single, cathodal band of species-specific ADH. During development at 26 degrees C, hybrid fry showed a preferential activation of the maternally derived Adh allele. It is suggested that the low activity of the paternally derived allele may result from an incompatibility between maternal regulatory factors and the paternal regulative element controlling gene expression.     

Transitional hemizygosity of the maternally derived allele at the 6PGD locus during early development of the Cyprinid fish Rutilus rutilus

The activation of individual alleles during early embryogenesis was studied at the 6-phosphogluconate dehydrogenase gene locus of the Cyprinid fish Rutilus by means of starch gel electrophoresis. By using three alleles occurring at this locus, it was possible to discriminate between (1) maternally transmitted gene products stored in the egg cytoplasm, (2) newly synthesized protein of the maternally derived allele in the embryonic genome, and (3) newly synthesized protein of the paternally derived allele. It was found that, until the fifth day of development, maternal products were present in the embryo. By the seventh day after fertilization, these storage products were nearly exhausted, and a hemizygous phenotype for the maternally derived gene became visible. On the eighth day, the patterns of all four allelic combinations of the mating type used were demonstrable in the offspring. The findings suggest that for the alleles used in this study, the maternally derived gene is preferentially activated during embryogenesis.Supported by the Deutsche Forschungsgemeinschaft.     

A fine analysis of glucose-phosphate-isomerase patterns in single preimplantation mouse embryos

Mouse embryos were derived from eggs heterozygous for alleles of the dimeric enzyme glucose phosphate isomerase (Gpi-1a/Gpi-1b) that had been fertilized with sperm carrying a third allele (Gpi-1c). This particular combination makes it possible to study the activity of the paternally derived as well as the maternally derived genes, the persistence of oocyte-coded enzyme throughout early development and the possible simultaneous expression of both the paternally derived allele and the maternal message. The different isozymes present in single embryos were separated by electrophoresis. The results show that the oocyte-coded glucose phosphate isomerase is gradually replaced by embryo-coded enzyme. Expression of the paternally derived allele was first detected at the morula stage, during which the translation of the maternally derived message seemed to be either exhausted or below the detection limit of our system. Some oocyte-coded enzyme persisted until after implantation.     

Expression of specific genes in early mouse embryos blocked by cytochalasin

Summary Mouse embryos at the two cell stage derived from C57BL/6 × C3H/Aa F₁-females heterozygous at the X-linked phosphoglycerate kinase locus (Pgk-1) were cultured continuously in the presence of cytochalasin B or D. Further cleavage of the two cell embryos was thus prevented and the embryos became polyploid during culture. The onset of expression of the maternally inherited Pgk-1 gene and of the paternally inherited glucosephosphate isomerase (Gpi-1) gene was determined in these polyploid embryos by cellulose acetate gel electrophoresis of single embryos. In contrast to euploid preimplantation embryos developing normally in utero or in culture without cytochalasins, expression of maternal Pgk-1 was never observed at days 4 and 5 of gestation in polyploid two cell embryos, showing that the Pgk-1 allele on the maternally inherited X chromosome is not activated independently of cytokinesis and morphogenesis. Expression of paternally derived Gpi-1, however, occurred in cleavage blocked embryos von day 5 of development. This may indicate that the activation of two genes which are both expressed during preimplantation development and which both code for glycolytic enzymes, is initiated by different signals.     

Association of an isozyme locus and strawbreaker foot rot resistance derived from Aegilops ventricosa in wheat

Summary Thirty lines from a cross between VPM/ Moisson 421 and Selection 101 were used in the study to determine whether strawbreaker foot rot resistance derived from Aegilops ventricosa was associated with an allele for endopeptidase. The progeny examined for foot rot lesions represented F₂ derived F₅ lines and enzyme assays were done on F₆ seedlings. The results indicate that the wheat and VPM/Moisson 421 endopeptidase alleles are distinctly different. The endopeptidase allele frequencies of 30 lines were compared with strawbreaker foot rot resistance as measured by the lesion severity index. The results demonstrate a close association between the gene for strawbreaker foot rot resistance and the endopeptidase allele derived from Ae. ventricosa.     

Parental influences on expression of glucose-6-phosphate dehydrogenase, G6pd, in the mouse; a case of imprinting

Glucose-6-phosphate dehydrogenase (G6PD) activity was measured in blood from heterozygotes for the normal allele G6pda and the low activity allele G6pda-mlNeu. In adult mice lower activity was found in G6pda/G6pda-mlNeu than in the reciprocal heterozygote G6pda-mlNeu/G6pda (the maternal allele being listed first). Thus, either the paternally derived allele was over-expressed or the maternally derived allele was under-expressed. By contrast, in younger mice the difference in G6PD activity in reciprocal crosses was less marked. The findings are interpreted in terms of differential imprinting of maternally and paternally inherited information. The explanation offered for age related differences is that, as a consequence of imprinting, either the paternal X-chromosome is preferentially reactivated, or cells in which the paternally derived allele is active are at a selective advantage, and proliferate better than those in which the maternally inherited allele is active.     

The Expression of X-Linked Phosphoglycerate Kinase in the Early Mouse Embryo

Abstract. During early mouse embryogenesis, the activity of X-chromosomally linked maternal and paternal phosphoglycerate kinase (PGK-1) alleles was determined using electrophoretic separation of their gene products and a sensitive fluorometric enzyme assay. In the embryos collected from females homozygous for PGK-1^b mated with PGK-1^a males and vice versa, the paternally derived allozyme was first detected after implantation on day 6. Expression of the maternally inherited allele was studied in embryos from females heterozygous for PGK-1^b and PGK-1^a. From day 1 to day 4, the embryos maintained a constant ratio of enzyme activity of PGK-1B to PGK-1A. Prior to implantation of the embryos between day 4 and day 5, the activity ratio of the two PGK-1 allelic variants changed significantly due to the first appearance of newly synthesized PGK derived from the maternally inherited allele.
Our data demonstrate a temporal difference in the onset of PGK synthesis depending on whether this particular gene product is of maternal or paternal origin. Therefore, we conclude that the maternal PGK-1 locus is already activated during late preimplantation development whereas the paternally inherited gene locus remains silent at the preimplantation stage but is subsequently expressed at approximately the time of X-chromosomal inactivation.     

Synchronous activation of both parental alleles at the 6-PGD locus of Japanese quail embryos

The gene locus for the enzyme 6-phosphogluconate dehydrogenase belongs to that part of the genome which is activated at the beginning of embryonic development. The present experiment, utilizing three alleles at this autosomally inherited locus of the Japanese quail, was designed to show whether exhaustion of maternally stored 6-PGD is followed by maternally hemizygous de novo synthesis of the same enzyme. 6-PGD phenotypes of early embryos resulting from the mating between a male homozygous for one allele and a female heterozygous for two other alleles were examined by starch gel electrophoresis. The result showed that the maternally stored 6-PGD is exhausted before the twenty-fourth hour of incubation. This is followed by synchronous activation of both parental alleles. Previous studies on the development of various interspecific crosses have revealed that, at all loci studied, the activation of the maternally derived allele preceded that of the paternally derived allele. The present experiment reveals that preferential activation of maternally derived alleles need not be a rule of development.This work was supported in part by a grant (CA-05138) from the National Cancer Institute, U.S. Public Health Service, and in part by a research fund established in honor of General James H. Doolittle. Contribution No. 6-68, Department of Biology, City of Hope Medical Center.     

Regulation of the activity of alcohol dehydrogenase isozymes during the development of the maize kernel

Summary The relative activities of alcohol dehydrogenase isozymes have been studied during the development of the endosperm and scutellum of heterozygous Adh ₁ ^F /Adh ₁ ^S maize kernels. The products of the Adh ₁ ^F allele are found earlier than the products of the Adh ₁ ^S allele in both the scutellum and the endosperm. A second gene (Adh _r )which controlsthe activity level of ADH is active in the scutellum only. The Adh _r ^N allele specifies increase in the relative activity of the Adh ₁ ^S products from 26 to 38 days after pollination. This increase is prevented by the Adh _r ^L allele which is dominant. These results ar discussed on the basis of the limited factor hypothesis proposed recently by Schwartz (1971) for the regulation of the Adh ₁gene in maize.     

Reproductive barrier and genomic imprinting in the endosperm of flowering plants

In flowering plants, success or failure of seed development is determined by various genetic mechanisms. During sexual reproduction, double fertilization produces the embryo and endosperm, which both contain maternally and paternally derived genomes. In endosperm, a reproductive barrier is often observed in inter-specific crosses. Endosperm is a tissue that provides nourishment for the embryo within the seed, in a similar fashion to the placenta of mammals, and for the young seedling after germination. This review considers the relationship between the reproductive barrier in endosperm and genomic imprinting. Genomic imprinting is an epigenetic mechanism that results in mono-allelic gene expression that is parent-of-origin dependent. In Arabidopsis, recent studies of several imprinted gene loci have identified the epigenetic mechanisms that determine genomic imprinting. A crucial feature of genomic imprinting is that the maternally and paternally derived imprinted genes must carry some form of differential mark, usually DNA methylation and/or histone modification. Although the epigenetic marks should be complementary on maternally and paternally imprinted genes within a single species, it is possible that neither the patterns of epigenetic marks nor expression of imprinted genes are the same in different species. Moreover, in hybrid endosperm, the regulation of expression of imprinted genes can be affected by upstream regulatory mechanisms in the male and female gametophytes. Species-specific variations in epigenetic marks, the copy number of imprinted genes, and the epigenetic regulation of imprinted genes in hybrids might all play a role in the reproductive barriers observed in the endosperm of interspecific and interploidy crosses. These predicted molecular mechanisms might be related to earlier models such as the "endosperm balance number" (EBN) and "polar nuclei activation" (PNA) hypotheses.     

X-Linked Gene Expression in the Virginia Opossum: Differences between the Paternally Derived Gpd and Pgk-A Loci

Expression of X-linked glucose-6-phosphate dehydrogenase (G6PD) and phosphoglycerate kinase-A (PGK-A) in the Virginia opossum (Didelphis virginiana) was studied electrophoretically in animals from natural populations and those produced through controlled laboratory crosses. Blood from most of the wild animals exhibited a common single-banded phenotype for both enzymes. Rare variant animals, regardless of sex, exhibited single-banded phenotypes different in mobility from the common mobility class of the respective enzyme. The laboratory crosses confirmed the allelic basis for the common and rare phenotypes. Transmission of PGK-A phenotypes followed the pattern of determinate (nonrandom) inactivation of the paternally derived Pgk-A allele, and transmission of G6PD also was consistent with this pattern. A survey of tissue-specific expression of G6PD phenotypes of heterozygous females revealed, in almost all tissues, three-banded patterns skewed in favor of the allele that was expressed in blood cells. Three-banded patterns were never observed in males or in putatively homozygous females. These patterns suggest simultaneous, but unequal, expression of the maternally and paternally derived Gpd alleles within individual cells (i.e., partial paternal allele expression). The absence of such partial expression was noted in a parallel survey of females heterozygous at the Pgk-A locus. Thus, it appears that Gpd and Pgk-A are X-linked in D. virginiana and subject to preferential paternal allele inactivation, but that dosage compensation may not be complete for all paternally derived X-linked genes. The data establish the similarity between the American and Australian marsupial patterns of X-linked gene regulation and, thus, support the hypothesis that this form of dosage compensation was present in the early marsupial lineage that gave rise to these modern marsupial divisions. In addition, the data provide the first documentation of the differential expression of two X-linked genes in a single marsupial species. Because of its combination of X-linked variation, high fecundity, and short generation time, D. virginiana is a unique model for pursuing questions about marsupial gene regulation that have been difficult to approach through studies of Australian species.     

Autonomous endosperm development in flowering plants: how to overcome the imprinting problem?

In the vast majority of sexually reproducing flowering plants, a ratio of 2 maternally derived genomes to 1 paternally derived genome (2m:1p) is essential for normal endosperm development, and therefore ultimately for seed development. Even in many pseudogamous apomicts, where the embryo develops without a paternal contribution, fertilisation of the endosperm to obtain the correct 2m:1p parental ratio is still necessary. How do autonomous apomicts, where both embryo and endosperm develop autonomously, circumvent this requirement? The background for the 2m:1p requirement is that the parental genomes are epigenetically different; in either genome, a set of genes is silenced in a sex-specific way by genomic imprinting. Removal of the imprints from the maternally derived endosperm genome leads to expression of normally maternally silenced genes, and effectively supplies the missing paternal genome. In Arabidopsis, we propose that a combination of the fie mutation and hypomethylation of the genome creates such a situation in the endosperm genome. As a result, in a fie mutant, hypomethylated ovule complete autonomous endosperm development takes place in the absence of fertilisation.     

Allelic inhibition at the autosomally inherited gene locus for liver alcohol dehydrogenase in chicken-quail hybrids

At the gene locus for liver alcohol dehydrogenase (ADH) of the Japanese quail, three alleles which give electrophoretic variants, A, B, and C, exist. This enzyme is autosomally inherited. Allelic polymorphism was not observed in the chicken, but the wild-type ADH of the chicken can readily be distinguished from A, B, and C of the quail by starch gel electrophoresis. In the development of both species, ADH activity reached a near adult level at about the nineteenth day (a few days after hatching in the quail and a few days before hatching in the chicken). Chicken-quail hybrids at the day of hatching (nineteenth day) revealed the presence of maternally derived quail ADH only, and their ADH activities were about half that of both parental species. Those hybrids which received either A or C allele from the mother quail showed three bands of ADH at the third day after hatching. The chicken and quail alleles began to function in synchronous harmony. One 3-day-old and two adult hybrids which received B allele from the quail, however, still revealed complete absence of the paternally derived chicken ADH.This work was supported in part by a grant (CA-05138) from the National Cancer Institute, U.S. Public Health Service, and in part by a research fund established in honor of General James H. Doolittle. Contribution No. 20-67, Department of Biology, City of Hope Medical Center.Dr. Eduardo Castro-Sierra is a fellow of the Institute for Advanced Learning of the City of Hope Medical Center.