Role of hypermutability in the evolution of the genus Oenococcus

Oenococcus oeni is an alcohol-tolerant, acidophilic lactic acid bacterium primarily responsible for malolactic fermentation in wine. A recent comparative genomic analysis of O. oeni PSU-1 with other sequenced lactic acid bacteria indicates that PSU-1 lacks the mismatch repair (MMR) genes mutS and mutL. Consistent with the lack of MMR, mutation rates for O. oeni PSU-1 and a second oenococcal species, O. kitaharae, were higher than those observed for neighboring taxa, Pediococcus pentosaceus and Leuconostoc mesenteroides. Sequence analysis of the rpoB mutations in rifampin-resistant strains from both oenococcal species revealed a high percentage of transition mutations, a result indicative of the lack of MMR. An analysis of common alleles in the two sequenced O. oeni strains, PSU-1 and BAA-1163, also revealed a significantly higher level of transition substitutions than were observed in other Lactobacillales species. These results suggest that the genus Oenococcus is hypermutable due to the loss of mutS and mutL, which occurred with the divergence away from the neighboring Leuconostoc branch. The hypermutable status of the genus Oenococcus explains the observed high level of allelic polymorphism among known O. oeni isolates and likely contributed to the unique adaptation of this genus to acidic and alcoholic environments.     

A study of the mechanism of the antiarrhythmic action of Allapinin

Allapinin (lappaconitine hydrobromide) is a drug used for the treatment of cardiac rhythm disturbances; its properties are characteristic of class IC antiarrhythmics. The mechanism of its electrophysiological action involves the blockade of Na⁺ channels with a subsequent decrease of depolarization rate leading to a slowing of impulse propagation and a decrease of excitability in the conductive system of the heart. Factors underlying the side effects of Allapinin (tachycardia, arterial hypertension, impaired coordination, etc.) are currently unknown, and therefore a study of the molecular mechanisms of its action seems relevant. The target genes of the drug were identified in rats with induced aconitine arrhythmia using the commercially available Rat Neuroscience Ion Channels & Transporters RT² Profiler? PCR Array kit (SA Biosciences). A comparison of expression levels of 84 genes in rats treated with Allapinin, after the induction of arrhythmia by aconitine (experiment) and in physiological saline-treated arrhythmic rats (control), revealed 18 mRNAs which were up- or downregulated twofold or more in the experiment relative to the control. Allapinin was shown to stimulate the expression of genes coding for various types of K⁺ channels (kcna6, kcnj1, kcnj4, kcnq2, and kcnq4), Ca²⁺ channel (cacna1g), and vesicular acetylcholine transporter (slc18a3). A decrease in mRNA levels was detected for genes coding for K⁺ channels (kcne1, kcns1), a Na⁺ channel (scn8a), and membrane transporter genes (atp4a, slc6a9). Our data shows that Allapinin administered to animals with aconitine arrhythmia modulates the expression of genes accounting for ion current conductances involved in the formation of various phases of action potential (I _Na , I _to , I _Ks , I _K1 , I _CaT ). The effect of the drug on the levels of mRNAs coding for acetylcholine and glycine transporters suggests the involvement of these neuromediators in the mechanisms underlying the antiarrhythmic effect of Allapinin.     

Effect of atoms evaporated from the anode on the structure of the vacuum arc space charge sheath

The structure of the anode space charge sheath of a vacuum arc is studied with allowance for the dependence of the negative anode fall on the ratio of the directed electron velocity v ₀ to the electron thermal velocity v _T for different values of the flux density of atoms evaporated from the anode. Poisson’s equation for the sheath potential is solved taking into account the electron space charge, fast cathode ions, and slow ions produced due to the ionization of atoms evaporated from the anode. The kinetic equation for atoms and slow anode ions is solved with allowance for ionization in the collision integral. Analytic solutions for the velocity distribution functions of atoms and slow ions and the density of slow ions are obtained. It is shown that the flux of slow ions substantially affects the spatial distribution of the electric field E(z) in the sheath. As the flux density increases, the nonmonotonic dependence E(z) transforms into a monotonic one and the sheath narrows. For a given flux of evaporated atoms Πa, the increase in the ratio of the directed electron velocity to the electron thermal velocity leads again to a nonmonotonic dependence E(z). As z increases, the electric field first increases, passes through the maximum, decreases, passes through the minimum E _min, and then again increases toward the anode. There is a limiting value of the ratio (v ₀/v _T )* at which E _min(z) vanishes. At v ₀/v _T > (v ₀/V _T )*, the condition for the existence of a steady-state sheath is violated and the profiles of the field and potential in the sheath become oscillating. The dependence of (v ₀/v _T )* on the flux density of evaporated atoms Π _a is obtained. It is shown that the domain of existence of steady-state solutions in the sheath broadens with increasing Π _a .     

Role of the parCBA Operon of the Broad-Host-Range Plasmid RK2 in Stable Plasmid Maintenance

The par region of the stably maintained broad-host-range plasmid RK2 is organized as two divergent operons, parCBA and parDE, and a cis-acting site. parDE encodes a postsegregational killing system, and parCBA encodes a resolvase (ParA), a nuclease (ParB), and a protein of unknown function (ParC). The present study was undertaken to further delineate the role of the parCBA region in the stable maintenance of RK2 by first introducing precise deletions in the three genes and then assessing the abilities of the different constructs to stabilize RK2 in three strains of Escherichia coli and two strains of Pseudomonas aeruginosa. The intact parCBA operon was effective in stabilizing a conjugation-defective RK2 derivative in E. coli MC1061K and RR1 but was relatively ineffective in E. coli MV10Δlac. In the two strains in which the parCBA operon was effective, deletions in parB, parC, or both parB and parC caused an approximately twofold reduction in the stabilizing ability of the operon, while a deletion in the parA gene resulted in a much greater loss of parCBA activity. For P. aeruginosa PAO1161Rif^r, the parCBA operon provided little if any plasmid stability, but for P. aeruginosa PAC452Rif^r, the RK2 plasmid was stabilized to a substantial extent by parCBA. With this latter strain, parA and res alone were sufficient for stabilization. The cer resolvase system of plasmid ColE1 and the loxP/Cre system of plasmid P1 were tested in comparison with the parCBA operon. We found that, not unlike what was previously observed with MC1061K, cer failed to stabilize the RK2 plasmid with par deletions in strain MV10Δlac, but this multimer resolution system was effective in stabilizing the plasmid in strain RR1. The loxP/Cre system, on the other hand, was very effective in stabilizing the plasmid in all three E. coli strains. These observations indicate that the parA gene, along with its res site, exhibits a significant level of plasmid stabilization in the absence of the parC and parB genes but that in at least one E. coli strain, all three genes are required for maximum stabilization. It cannot be determined from these results whether or not the stabilization effects seen with parCBA or the cer and loxP/Cre systems are strictly due to a reduction in the level of RK2 dimers and an increase in the number of plasmid monomer units or if these systems play a role in a more complex process of plasmid stabilization that requires as an essential step the resolution of plasmid dimers.     

Checklist of the species of the aseptate Gregarine family Urosporidae

A checklist is given of the 45 named species of the family Urosporidae (phylum Protozoa, subphylum Apicomplexa, class Sporozoa, subclass Gregarinia, order Eugregarinida, suborder Aseptatina) together with their synonyms, the names of their hosts, their locations in the hosts, their known geographic distribution, and key references. Another list is given of synonyms, lapsi calami, nomina nuda, etc. associated with the genera of this family. The following taxonomic-nomenclatural innovations are introduced—NEW NAME: Gonospora good-richae nom. nov. in the polychaete Arenicola ecaudata; NEW COMBINATIONS: Urospora grassei (Changeux, 1961) in the sea cucumbers Holothuria spp.; Urospora schneideri (Mingazzini, 1891) in the sea cucumbers Holothuria spp; Gonospora gonadipertha (Djakonov, 1923) in the sea cucumber Cucumaria frondosa; Gonospora stichopi (Lützen, 1967) in the sea cucumber Stichopus tremulus.     

Evidence for AEGILOPS SHARONENSIS Eig as the Donor of the B Genome of Wheat

A number of lines of evidence are advanced for the candidacy of Aegilops sharonensis Eig as the donor of the B genome of wheat. The cytoplasm of Ae. sharonensis is compatible with tetraploid wheat Triticum turgidum dicoccoides, as evidenced by the high level of chromosome pairing and fertility of the amphiploid Ae. sharonensis x T. turgidum dicoccoides. Ae. sharonensis chromosomes exhibit high levels of pairing with those of the B genome of wheat in hybrids with Ph-deficient hexaploid wheat and low levels of homoeologous pairing with T. monococcum chromosomes.——The amphidiploid between Ae. sharonensis and T. monococcum is very similar to T. turgidum dicoccoides in spike, spikelet and grain morphology. The karyotype of Ae. sharonensis resembles more closely that of extrapolated B genome karyotypes of wheat than do the karyotypes of other proposed B-genome donor species of Aegilops. Because of distinctiveness in cytological affinity and karyotype morphology between Ae. sharonensis and Ae. longissima, a separate genome symbol S^sh is proposed for the former species.     

Studies on the interactions of sperm with the surface of the sea urchin egg

We have examined the relationship between sperm adhesion and fertilization in the cross species insemination of Arbacia punctulata eggs by Strongylocentrotus purpuratus sperm. As previously reported (Kinsey et al., 1980) the addition of S. purpuratus egg jelly results in induction of the acrosome reaction in sperm and significant numbers of S. purpuratus sperm adhere to A. punctulata eggs. However, in the absence of S. purpuratus egg jelly, S. purpuratus sperm fail to bind to A. punctulata eggs. Although at least 200 S. purpuratus sperm bind to an A. punctulata egg in the presence of S. purpuratus jelly, less than 8% of the eggs are fertilized. The adhesion of S. purpuratus sperm meets the same functional criteria as homologous A. punctulata sperm-egg adhesion. Electron microscopy shows that S. purpuratus sperm that have undergone the acrosome reaction adhere to A. punctulata eggs by their bindin-coated acrosomal process in a manner that is morphologically identical to that observed with homologous A. punctulata sperm. We have also compared the ability of S. purpuratus and A. punctulata sperm to fuse and fertilize with A. punctulata eggs after removal of the vitelline layer. Using high levels of sperm of either species, heterologous as well as homologous fertilization is readily detectable. Under these conditions, where stable binding is not demonstrable, there is no difference in the ability of S. purpuratus and A. punctulata sperm to fertilize A. punctulata eggs. These observations suggest that the failure of S. purpuratus sperm to fertilize A. punctulata eggs under normal conditions may be due to their inability to penetrate the vitelline layer so that they can fuse with the egg plasma membrane. In relation to the possible mechanism of vitelline layer penetration, we have also investigated the mode of action of chymostatin, an inhibitor of chymotrypsin that has been reported to inhibit fertilization of sea urchin eggs (Hoshi et al., 1979). Our findings suggest that the fertilization inhibitory activity of chymostatin is not related to its antichymotrypsin activity. Rather, it appears that this inhibition is due to the induction of an abnormal acrosome reaction in sperm that precludes formation of the acrosome process.     

Cloning and Characterization of the Gene Cluster Involved in the Production of the Circular Bacteriocin Carnocyclin A

Carnocyclin A is a circular bacteriocin of 60 amino acids produced by Carnobacterium maltaromaticum UAL307. A region of 12 kb that contained the structural gene for carnocyclin A, cclA, was sequenced using a fosmid library, and 10 genes were identified that could be responsible for carnocyclin A production and immunity. Five of those genes, cclBITCD, were found upstream of cclA: one encodes a protein containing a conserved ATP-binding domain and four encode proteins with putative membrane-spanning domains. CclC shows homology with a family of membrane proteins that contain the domain of unknown function 95 (DUF95). Downstream of cclA four additional genes, cclEFGH, were identified that show similarity to the last four genes, as-48EFGH, of the enterocin AS-48 bacteriocin gene cluster. CclFGH shows sequence homology with As-48FGH. Transformation of C. maltaromaticum UAL26 with cclBITCDA resulted in production of carnocyclin A, indicating that these genes form the minimal requirement for the secretion of fully matured bacteriocin. cclI encodes for a small hydrophobic protein with a high pI, which are characteristic features of known immunity proteins for other circular bacteriocins. Indeed, cloning of cclI behind a constitutive promoter in UAL26 resulted in immunity although the level of resistance was lower than that of UAL26 containing cclBITCDA, indicating that CclI alone is not enough to confer full immunity to carnocyclin A.     

Testing the effect of biological control agents on the formation of vesicular arbuscular mycorrhiza

A container system for rapid infection of roots with pathogenic or mycorrhizal fungi was used to test the effect of the two commercial biological control agents, Trichoderma harzianum and Streptomyces griseoviridis, on the formation of vesicular-arbuscular mycorrhiza in soybean. In the presence of these biocontrol agents, mycorrhiza formation with Glomus mosseae was significantly depressed, particularly with S. griseoviridis. Infection by the root pathogen Rhizoctonia solani was not altered by these agents. Remarkably, not only R. solani but also T. harzianum induced accumulation of large amounts of the phytoalexin glyceollin in the roots. In contrast, roots inoculated with S. griseoviridis or with the mycorrhizal fungus G. mosseae did not accumulate glyceollin.     

Polyphasic approach to the taxonomy of the Rhizopus stolonifer group

Phylogenetic analysis of Rhizopus strains based on the D1/D2 region of LSU rDNA sequences yielded a phylogram with four well-supported clades. The R. microsporus clade concurs with classification obtained by traditional methods. The R. oryzae group was found to include species of the genus Amylomyces. The traditional R. stolonifer group was divided into two well-supported clades in the phylogram, with one clade comprising R. stolonifer var. stolonifer, R. sexualis var. sexualis, and R. sexualis var. americanus and the other clade comprising taxa with recurved sporangiophores; R. reflexus, R. stolonifer var. lyococcus, and R. circinans, identifying recurved sporangiophores as an important taxonomic character. The molecular data supported the recognition of this clade at the species level: R. lyococcus (basionym: Sporotrichum lyococcum).     

The role of recombination in the formation of circular oligomers of the lambda plasmid

The λdv1 plasmid forms an extensive oligomeric series of circular DNA molecules in recombination-proficient (recsu⁺) Escherichia coli. These rec⁺ [λdv1]⁺ strains can be typed into the following four classes according to which member of the oligomeric series is most frequent: monomer, dimer, trimer, and tetramer strains. Each of these strains forms a set of circular λdv1 DNA molecules in which most members belong to the series l, 2l, 3l, 4l, where l is the length of the most frequent circular DNA that characterizes the strain—i.e. l equals the length of the most frequent oligomer in the respective strain. In a given strain, the frequency of a molecular species decreases as its length becomes a larger multiple of l. For example, the dimer strains produce dimers, tetramers, hexamers, octomers, etc., in decreasing frequencies, which reach the limits of detection at about the hexadecamer.When recA^? mutations that are absolutely defective for host recombination are introduced into each of these four strains, l retains the same values as in the parent rec⁺ strain, but oligomers larger than 2l are not formed, and the frequency of the 2l oligomer is much reduced. The introduction of recB^? or recC^? mutations, which are only partially defective for host recombination, produces a much smaller perturbation of the rec⁺ distributions, and $\text{rec}{}^{\mathrm{+}}\text{recA}{}^{\mathrm{?}}$ merodiploids exhibit the rec⁺ phenotype with respect to both oligomerization and host recombination.The effects of rec^? mutations on the distribution of λdv1 oligomers and the nature of the oligomeric series produced in rec⁺ cells all indicate that an intermolecular reciprocal recombination between two circular λdv1 DNAs is the principal reaction responsible for oligomerization. It is suggested that the small residual oligomerization that yields 2l oligomers in recA^?cells results from aberrant segregation of the DNA strands at the termination of the replication of l-sized molecules.The inactivation of recA, but not of recB or C, also results in a marked reduction in the frequency of spontaneous curing which in recA⁺ [λdv1⁺]hosts leads to the segregation of [λdv^?]cells. However, spontaneous curing does not appear to be dependent upon the recombination reactions that yield the [λdv 1⁺]oligomers, since the frequency of oligomerization in recA⁺ hosts decreases with increasing l, whereas the frequency of curing increases with increasing l.     

Phylogenetic status of Xylaria subgenus Pseudoxylaria among taxa of the subfamily Xylarioideae (Xylariaceae) and phylogeny of the taxa involved in the subfamily

To infer the phylogenetic relationships of Xylaria species associated with termite nests within the genus Xylaria and among genera of the subfamily Xylarioideae, β-tubulin, RPB2, and α-actin sequences of 131 cultures of 114 species from Xylaria and 11 other genera of the subfamily were analyzed. These 11 genera included Astrocystis, Amphirosellinia, Discoxylaria, Entoleuca, Euepixylon, Kretzschmaria, Nemania, Podosordaria, Poronia, Rosellinia, and Stilbohypoxylon. We showed that Xylaria species were distributed among three major clades, TE, HY, and PO, with clade TE—an equivalent of the subgenus Pseudoxylaria—encompassing exclusively those species associated with termite nests and the other two clades containing those associated with substrates other than termite nests. Xylaria appears to be a paraphyletic genus, with most of the 11 genera submerged within it. Podosordaria and Poronia, which formed a distinct clade, apparently diverged from Xylaria and the other genera early. Species of Entoleuca, Euepixylon, Nemania, and Rosellinia constituted clade NR, a major clade sister to clade PO, while those of Kretzschmaria were inserted within clade HY and those of Astrocystis, Amphirosellinia, Discoxylaria, and Stilbohypoxylon were within clade PO.     

Changes in the Daily Rhythm of Lipid Metabolism in the Diabetic Retina

Disruption of circadian regulation was recently shown to cause diabetes and metabolic disease. We have previously demonstrated that retinal lipid metabolism contributed to the development of diabetic retinopathy. The goal of this study was to determine the effect of diabetes on circadian regulation of clock genes and lipid metabolism genes in the retina and retinal endothelial cells (REC). Diabetes had a pronounced inhibitory effect on the negative clock arm with lower amplitude of the period (per) 1 in the retina; lower amplitude and a phase shift of per2 in the liver; and a loss of cryptochrome (cry) 2 rhythmic pattern in suprachiasmatic nucleus (SCN). The positive clock arm was increased by diabetes with higher amplitude of circadian locomotor output cycles kaput (CLOCK) and brain and muscle aryl-hydrocarbon receptor nuclear translocator-like 1 (bmal1) and phase shift in bmal1 rhythmic oscillations in the retina; and higher bmal1 amplitude in the SCN. Peroxisome proliferator-activated receptor (PPAR) α exhibited rhythmic oscillation in retina and liver; PPARγ had lower amplitude in diabetic liver; sterol regulatory element-binding protein (srebp) 1c had higher amplitude in the retina but lower in the liver in STZ- induced diabetic animals. Both of Elongase (Elovl) 2 and Elovl4 had a rhythmic oscillation pattern in the control retina. Diabetic retinas lost Elovl4 rhythmic oscillation and had lower amplitude of Elovl2 oscillations. In line with the in vivo data, circadian expression levels of CLOCK, bmal1 and srebp1c had higher amplitude in rat REC (rREC) isolated from diabetic rats compared with control rats, while PPARγ and Elovl2 had lower amplitude in diabetic rREC. In conclusion, diabetes causes dysregulation of circadian expression of clock genes and the genes controlling lipid metabolism in the retina with potential implications for the development of diabetic retinopathy.     

Analysis and Expansion of the Role of the Escherichia coli Protein ProQ

The decrease in proline transport by the proline porter ProP in a ΔproQ strain has been well documented; however, the reason for this phenotype remains undefined. Previous studies have speculated that ProQ facilitates translation of proP mRNA. Here, we demonstrate that ProQ is enriched in the polysome fractions of sucrose gradient separations of E. coli lysates and the 30S fractions of lysates separated under conditions causing ribosomal subunit dissociation. Thus, ProQ is a bona fide ribosome associated protein. Analysis of proQ constructs lacking predicted structural domains implicates the N-terminal domain in ribosome association. Association with the ribosome appears to be mediated by an interaction with the mRNA being translated, as limited treatment of lysates with Micrococcal Nuclease maintains ribosome integrity but disrupts ProQ localization with polysomes. ProQ also fails to robustly bind to mRNA-free 70S ribosomes in vitro. Interestingly, deletion of proP does not disrupt the localization of ProQ with translating ribosomes, and deletion of proP in combination with the proU operon has no effect on ProQ localization. We also demonstrate that ProQ is necessary for robust biofilm formation, and this phenotype is independent of ProP. Binding studies were carried out using tryptophan fluorescence and in vitro transcribed proP mRNAs. proP is transcribed from two differentially regulated promoters, and ProQ interacts with proP mRNA transcribed from both promoters, as well as a control mRNA with similar affinities. In total, these data suggest that ProQ is positioned to function as a novel translational regulator, and its cellular role extends beyond its effects on proline uptake by ProP.     

Role of Bacterial Exopolysaccharides (EPS) in the Fate of the Oil Released during the Deepwater Horizon Oil Spill

Halomonas species are recognized for producing exopolysaccharides (EPS) exhibiting amphiphilic properties that allow these macromolecules to interface with hydrophobic substrates, such as hydrocarbons. There remains a paucity of knowledge, however, on the potential of Halomonas EPS to influence the biodegradation of hydrocarbons. In this study, the well-characterized amphiphilic EPS produced by Halomonas species strain TG39 was shown to effectively increase the solubilization of aromatic hydrocarbons and enhance their biodegradation by an indigenous microbial community from oil-contaminated surface waters collected during the active phase of the Deepwater Horizon oil spill. Three Halomonas strains were isolated from the Deepwater Horizon site, all of which produced EPS with excellent emulsifying qualities and shared high (97-100%) 16S rRNA sequence identity with strain TG39 and other EPS-producing Halomonas strains. Analysis of pyrosequence data from surface water samples collected during the spill revealed several distinct Halomonas phylotypes, of which some shared a high sequence identity (≥97%) to strain TG39 and the Gulf spill isolates. Other bacterial groups comprising members with well-characterized EPS-producing qualities, such as Alteromonas , Colwellia and Pseudoalteromonas , were also found enriched in surface waters, suggesting that the total pool of EPS in the Gulf during the spill may have been supplemented by these organisms. Roller bottle incubations with one of the Halomonas isolates from the Deepwater Horizon spill site demonstrated its ability to effectively produce oil aggregates and emulsify the oil. The enrichment of EPS-producing bacteria during the spill coupled with their capacity to produce amphiphilic EPS is likely to have contributed to the ultimate removal of the oil and to the formation of oil aggregates, which were a dominant feature observed in contaminated surface waters.     

Molecular Systematics and Evolution of the Growth Hormone Introns in the Salmoninae

DNA sequence data was collected for the C and D introns in the duplicate growth hormone loci (GH1 and GH2) from Brachmystax lenok, two subspecies of Hucho hucho, Hucho (Parahucho) perryi, Salmo salar, Salmo trutta, Acantholingua ohridana (Salmothymus), six species of Salvelinus, eight species of Oncorhynchus including O. masou, and three outgroups including Thymallus thymallus, Coregonus artedi, and Coregonus clupeaformis. Phylogenetic analyses were performed using maximum parsimony and maximum likelihood (PAUP, version 4.08beta) with gaps as missing data and as a fifth base. B. lenok was basal in all of the trees and all of the other genera were monophyletic with the exception that A. ohridana always placed within Salmo, and H. hucho sp. often placed with B. lenok. The GH1 introns supported a sister relationship between Oncorhynchus and Salvelinus, while the combined GH2 introns were ambiguous at this node. This result contrasts with trees based on morphology and the ribosomal ITS1 sequences that support a sister relationship between Salmo and Oncorhynchus. The only estrogen response element (ERE) in the gene is found in the C intron and has mutated in GH2 in all of the species except B. lenok. The ERE element in GH1 has undergone another mutation in all of the species except for B. lenok, and members of the two genera Salvelinus and Oncorhynchus. Thus these latter two genera are the only ones with a difference in expression of GH1 and GH2 in the presence of estrogen. Differences in selective pressure on the introns in the duplicate genes in different taxa could account for the conflicting results obtained in the phylogenetic analysis.     

Functional interactions between the gene tetanic and the Shaker gene complex of Drosophila

Different phenotypes associated with the tetanic (tta) mutation such as appendage contraction, maternal effect and low viability and fertility are enhanced by one extra dose of the Shaker gene complex (ShC). The tta mutation is lethal with two extra doses of ShC. In addition, tta embryos have a defective nervous system. In this paper, I analyse the interaction between tta and ShC to gain insight into their relationship. Aneuploid analysis suggests that the lethality is due to an interaction of the tta mutation with the maternal effect (ME) region of this gene complex. Mutations in the ME region of ShC partially suppress this interaction. Trans-heterozygous combinations of MEI[l(1)305] and MEIII [l(1)459] mutations causes dominant lethality in a tta background. Trans-heterozygous combinations of an MEII [l(1)1359] mutation with the cited MEI and MEIII mutations are lethal in a tta background. Double mutant combinations and gene dosage experiments, suggest that tta also interacts with the viable (V) region of ShC. These specific genetic interactions indicate that tta and the ME and V regions of ShC are functionally related. These results, together with the previous electrophysiological, molecular and biochemical studies on these mutants suggest an interaction at the protein level. Thus, in the case of the V region, the tta gene product may modulate the activity of the K⁺ channels encoded in this region. Furthermore, the extreme dosage sensitivity of the interaction between tta and ShC suggests a stoichiometric requirement for the different gene products involved, which might be physically associated and form heteromultimers.     

Antagonistic Gene Activities Determine the Formation of Pattern Elements along the Mediolateral Axis of the Arabidopsis Fruit

The Arabidopsis fruit mainly consists of a mature ovary that shows three well defined territories that are pattern elements along the mediolateral axis: the replum, located at the medial plane of the flower, and the valve and the valve margin, both of lateral nature. JAG/FIL activity, which includes the combined functions of JAGGED (JAG), FILAMENTOUS FLOWER (FIL), and YABBY3 (YAB3), contributes to the formation of the two lateral pattern elements, whereas the cooperating genes BREVIPEDICELLUS (BP) and REPLUMLESS (RPL) promote replum development. A recent model to explain pattern formation along the mediolateral axis hypothesizes that JAG/FIL activity and BP/RPL function as antagonistic lateral and medial factors, respectively, which tend to repress each other. In this work, we demonstrate the existence of mutual exclusion mechanisms between both kinds of factors, and how this determines the formation and size of the three territories. Medial factors autonomously constrain lateral factors so that they only express outside the replum, and lateral factors negatively regulate the medially expressed BP gene in a non-autonomous fashion to ensure correct replum development. We also have found that ASYMMETRIC LEAVES1 (AS1), previously shown to repress BP both in leaves and ovaries, collaborates with JAG/FIL activity, preventing its repression by BP and showing synergistic interactions with JAG/FIL activity genes. Therefore AS gene function (the function of the interacting genes AS1 and AS2) has been incorporated in the model as a new lateral factor. Our model of antagonistic factors provides explanation for mutant fruit phenotypes in Arabidopsis and also may help to understand natural variation of fruit shape in Brassicaceae and other species, since subtle changes in gene expression may cause conspicuous changes in the size of the different tissue types.