Nucleotide sequence analysis of a novel globulin1 null allele from the Illinois high protein strain of maize

The highly polymorphic maize globulin1 (glbl) gene encodes an abundant embryo storage protein. The present study extends the analysis of glbl variants to further explore the nature of polymorphism at this locus. The null allele Glb1-N1Hb, derived from the Illinois High Protein (IHP) strain of maize was characterized at the molecular level by nucleotide sequence analysis. Among other differences, a single-base insertion leading to a premature termination codon in the carboxyl-terminal half of the otherwise normal protein was observed. The likely reasons for the absence of GLB1 protein accumulation in the IHP strain of maize are discussed.     

A tissue culture induced Adh1 null mutant of maize results from a single base change

Summary We have found a null Adh1 allele which arose as a somaclonal variant following tissue culture of maize embryos carrying Adh1-1S and Adh1-1F alleles. Cloning and sequencing shows that the mutant allele derives from Adh1-1S and that there has been a single base change in the coding region of the gene which converts and AAG lysine codon to a TAG stop codon. The rate of nucleotide substitution (two per 218 embryos cultured) is much greater than normal mutation rates.     

Alleles at the PRB3 locus coding for a disulfide-bonded human salivary proline-rich glycoprotein (Gl 8) and a null in an Ashkenazi Jew.

From electrophoretic analysis, we identified in the saliva of an Ashkenazi Jew a disulfide-bonded major glycoprotein variant (Gl 8) that is a product of the proline-rich protein (PRP) locus PRB3. A previous study of this variant protein misidentified it as Pa 2 and as a product of a different PRP locus. The other PRB3 allele in this individual is an apparent null. To identify the mutations, we sequenced the tandemly repetitious exon 3 (the major protein-coding portions) of both alleles. A CGT----TGT (Arg----Cys) mutation was found in one allele (PRB3Scys), which accounts for the disulfide-bonded and peroxidase-modifying properties of Gl 8. A single nucleotide insertion was found in the other allele (PRB3Mnull) that leads to a frameshift with a premature termination codon that causes an apparent lack of gene expression. Null alleles are frequent at PRP loci coding for basic and glycosylated PRPs, and the mechanism described might explain other null phenotypes among PRPs. From nucleotide comparisons, a model of intragenic unequal crossing-over is proposed to explain, in part, the generation of the PRB3Mnull allele. The Gl 8 protein variant is found in Ashkenazi Jews (gene frequency around .008) but not in the general white, black, or Japanese populations. It is interesting that products of different PRP genes, Gl 8 from PRB3 and Pa 1 from PRH1, are both disulfide bonded and probably modify salivary peroxidase (part of an important intraoral antibacterial system) through formation of disulfide-bonded heterodimers.     

Characterization of embryo globulins encoded by the maizeGlb genes

Two of the most abundant proteins in maize embryos are saline-soluble, water-insoluble globulins. One is aM _r 63,000 protein encoded by theGlbl gene and the other is aM _r 45,000 component encoded by theGlb2 gene. Both proteins accumulate to high levels during embryo development and are rapidly degraded during the early stages of seed germination. Amino acid composition analysis indicates that these proteins may serve as storage reserves to provide sources of nitrogen and carbon to the germinating embryo. Amino-terminal sequence analysis of the finalGlb1 gene product, GLB1, and its immediate precursor, GLB1′, indicates that the latter is proteolytically cleaved near the amino terminus to form GLB1. In addition to these biochemical studies, we describe the identification of a novel maize variant which lacks the protein product of theGlb2 gene. This contribution from the University of Illinois Agricultural Experiment Station was supported by grants from The Standard Oil Corporation, a wholly owned subsidiary of BP America, Inc., and the U.S. Department of Agriculture (No. 88-37262-3427).     

NKS1, Na(+)- and K(+)-sensitive 1, regulates ion homeostasis in an SOS-independent pathway in Arabidopsis

An Arabidopsis thaliana mutant, nks1-1, exhibiting enhanced sensitivity to NaCl was identified in a screen of a T-DNA insertion population in the genetic background of Col-0 gl1sos3-1. Analysis of the genome sequence in the region flanking the T-DNA left border indicated two closely linked mutations in the gene encoded at locus At4g30996. A second allele, nks1-2, was obtained from the Arabidopsis Biological Resource Center. NKS1 mRNA was detected in all parts of wild-type plants but was not detected in plants of either mutant, indicating inactivation by the mutations. Both mutations in NKS1 were associated with increased sensitivity to NaCl and KCl, but not to LiCl or mannitol. NaCl sensitivity was associated with nks1 mutations in Arabidopsis lines expressing either wild type or null alleles of SOS1, SOS2 or SOS3. The NaCl-sensitive phenotype of the nks1-2 mutant was complemented by expression of a full-length NKS1 allele from the CaMV35S promoter. When grown in medium containing NaCl, nks1 mutants accumulated more Na(+) than wild type and K(+)/Na(+) homeostasis was perturbed. It is proposed NKS1, a plant-specific gene encoding a 19kDa endomembrane-localized protein of unknown function, is part of an ion homeostasis regulation pathway that is independent of the SOS pathway.     

How Often Do Duplicated Genes Evolve New Functions?

A recently duplicated gene can either fix a null allele (becoming a pseudogene) or fix an (advantageous) allele giving a slightly different function, starting it on the road to evolving a new function. Here we examine the relative probabilities of these two events under a simple model. Null alleles are assumed to be neutral; linkage effects are ignored, as are unequal crossing over and gene conversion. These assumptions likely make our results underestimates for the probability that an advantageous allele is fixed first. When new advantageous mutations are additive with selection coefficient s and the ratio of advantageous to null mutations is ρ, the probability an advantageous allele is fixed first is ([1 - e(-S)]/[ρS] + 1)(-1), where S = 4N(e)s with N(e) the effective population size. The probability that a duplicate locus becomes a pseudogene, as opposed to evolving a new gene function, is high unless ρS & 1. However, even if advantageous mutations are very rare relative to null mutations, for sufficiently large populations ρS & 1 and new gene function, rather than pseudogene formation, is the expected fate of most duplicated genes.     

Abscisic Acid and the developmental regulation of embryo storage proteins in maize

The relationship of abscisic acid (ABA) inhibition of precocious germination and ABA-induced storage protein accumulation was examined over the course of embryogenesis in wild-type and viviparous mutants of maize (Zea mays L.). We show that a high level of embryo ABA and the product of the Viviparous-1 gene are both required in early maturation phase for germination suppression and the accumulation of storage globulins encoded by the gene Glb1. Suppressing precocious germination with a high osmoticum is not sufficient to initiate Glb1 protein synthesis, although continued accumulation is contingent upon this inhibition; germination of immature or mature embryos leads to a decline in synthesis and the degradation of stored globulins. Late in embryogenesis, fragments of Glb1 protein accumulate, coinciding with the loss of ABA sensitivity. These results suggest that ABA influences storage globulin accumulation by initiating synthesis, suppressing degradation, and inhibiting precocious germination.     

Experimental verification of microsatellite null alleles in Norway spruce (Picea abies [L.] Karst.): Implications for population genetic studies

Three stands ofPicea abies [L.] Karst. with different density in the Harz Mountains (Lower Saxony, Germany) were characterized at 4 microsatellite loci. An excess of homozygotes was observed in all 3 stands at 1 simple sequence repeat (SSR) locus, suggesting the presence of null alleles. To test for the segregation of a null allele, 24 openpollinated seeds (haploid megagametophytes and embryos) from apparently homozygous mother trees were analyzed. For 1 of 3 trees that could be identified as heterozygous for a null allele, no significant deviation from the expected 1∶1 segregation into marker absence (null allele) and marker presence of the second maternal allele could be observed in the haploid megagametophyte. Concordantly, the numbers of embryos heterozygous for the null allele and for the other maternal allele were not significantly different from each other. Inheritance analyses in seedlings and corresponding megagametophytes of gymnosperms were used as a direct experimental verification of microsatellite null alleles in single-tree progeny. Microsatellites with an abundance of null alleles should be discarded from further analysis because inclusion of these loci results in incorrect estimation of allele frequencies.     

Molecular relationships between alcohol dehydrogenase null-activity alleles from natural populations of Drosophila melanogaster.

Alcohol dehydrogenase null-activity alleles extracted from a number of natural populations of Drosophila melanogaster in Tasmania were shown to be molecularly similar by probing, with an oligonucleotide specific to an inserted region in intron 2 of the gene, genomic DNA amplified by the polymerase chain reaction. This insertion had previously been shown to be the cause of the loss of activity in one of the null alleles whose DNA sequence was known. Three Adh null alleles from mainland populations did not contain the insertion. Two of these null alleles, extracted from the Coffs Harbour population in different years, were cloned, and their DNA sequences showed that they were identical and that both had a 438-bp deletion which removed most of exon 2. The third null allele, identified in a sample of flies from Chateau Tahbilk, was shown by 4-bp restriction-endonuclease mapping to contain a 320-bp insertion in intron 1, although this may not be the cause of the loss of activity. The data show that at least three different Adh null alleles have been found in Australian populations and that at least two have been maintained as heterozygotes over a period of years.