Chiasma formation in Arabidopsis thaliana accession Wassileskija and in two meiotic mutants

Meiotic chiasmata were analysed in metaphase I pollen mother cells (PMCs) of wild-type Arabidopsis thaliana and in two meiotic mutants. Fluorescence in situ hybridisation (FISH) with 45S rDNA and 5S rDNA as probes was used to identify the five chromosome pairs. A wild-type chiasma frequency of 9.24 per cell was found, consistent with estimated genetic recombination values. Individual bivalent chiasma frequencies varied according to chromosome size; chromosome 1 had the highest mean chiasma frequency (2.14) while the short acrocentric chromosomes had the lowest frequencies (1.54 and 1.56). FISH analysis was extended to two meiotic mutants (asy1 and dsy1) having low residual bivalent and chiasma frequencies. Mutant dsy1 gave no indication of chromosome preference for residual bivalent formation; instead it showed a general reduction in bivalent and chiasma frequencies. In asy1, the longest chromosome (1) had the lowest bivalent frequency and chiasma frequency while the short acrocentric chromosome 2 had the highest frequencies. This chromosome pair may be preferentially involved in synapsis and chiasma formation because of their association with the nucleolus. However, other factors may be operating since the other acrocentric chromosome (4), with similar size and structure to chromosome 2, did not share these chiasma properties.     

A diverse and evolutionarily fluid set of microRNAs in Arabidopsis thaliana

To better understand the diversity of small silencing RNAs expressed in plants, we employed high-throughput pyrosequencing to obtain 887,000 reads corresponding to Arabidopsis thaliana small RNAs. They represented 340,000 unique sequences, a substantially greater diversity than previously obtained in any species. Most of the small RNAs had the properties of heterochromatic small interfering RNAs (siRNAs) associated with DNA silencing in that they were preferentially 24 nucleotides long and mapped to intergenic regions. Their density was greatest in the proximal and distal pericentromeric regions, with only a slightly preferential propensity to match repetitive elements. Also present were 38 newly identified microRNAs (miRNAs) and dozens of other plausible candidates. One miRNA mapped within an intron of DICER-LIKE 1 (DCL1), suggesting a second homeostatic autoregulatory mechanism for DCL1 expression; another defined the phase for siRNAs deriving from a newly identified trans-acting siRNA gene (TAS4); and two depended on DCL4 rather than DCL1 for their accumulation, indicating a second pathway for miRNA biogenesis in plants. More generally, our results revealed the existence of a layer of miRNA-based control beyond that found previously that is evolutionarily much more fluid, employing many newly emergent and diverse miRNAs, each expressed in specialized tissues or at low levels under standard growth conditions.     

Very-long-chain fatty acids have an essential role in plastid division by controlling Z-ring formation in Arabidopsis thaliana

Recent studies have showed the essential mechanisms for plastid division that have bacterial orthologues, such as FtsZ and Min system proteins; however, causal factors regulating plastid division in plant cells are poorly understood. Here, we show that plastid division is inhibited in Arabidopsis by reduced amounts of very-long-chain fatty acids (VLCFAs), which have an acyl chain length of more than 20 carbons and are used for cuticular wax formation. The number of amyloplasts and chloroplasts decreased in the mutant defective in VLCFA synthesis and in wild-type plants treated with an inhibitor of VLCFA synthesis. Although similar inhibition of plastid division was observed in transgenic plants that over-expressed PDV2 , one of the outer membrane proteins at the plastid division site, dot-like aggregates of FtsZ protein and disordered placement of multiple Z-rings were found in wild-type chloroplasts treated the inhibitor of VLCFA synthesis. Expression analysis showed that ARC3 , one of the Min system genes, was down-regulated under low VLCFA conditions. Our results indicate that VLCFAs or VLCFA-containing lipids have an essential role in plastid division by controlling Z-ring formation, showing a novel function of plant VLCFAs.     

Survival of Hoxa13 homozygous mutants reveals a novel role in digit patterning and appendicular skeletal development

The loss of HOXA13 function severely disrupts embryonic limb development. However, because embryos lacking HOXA13 die by mid‐gestation, the defects present in the mutant limb could arise as a secondary consequence of failing embryonic health. In our analysis of the mutant Hoxa13^GFP allele, we identified a surviving cohort of homozygous mutants exhibiting severe limb defects including: missing phalanx elements, fusions of the carpal/tarsal elements, and significant reductions in metacarpal/metatarsal length. Characterization of prochondrogenic genes in the affected carpal/tarsal regions revealed significant reduction in Gdf5 expression, whereas Bmp2 expression was significantly elevated. Analysis of Gdf5 mRNA localization also revealed diffuse expression in the carpal/tarsal anlagen, suggesting a role for HOXA13 in the organization of the cells necessary to delineate individual carpal/tarsal elements. Together these results identify Gdf5 as a potential target gene of HOXA13 target gene and confirm a specific role for HOXA13 during appendicular skeletal development. Developmental Dynamics 239:446–457, 2010. © 2009 Wiley‐Liss, Inc.     

Composition and formation of heterochromatin in Arabidopsis thaliana

The term heterochromatin has been applied to both large-scale, microscopically visible chromocentres and small-scale, silent genes located outside chromocentres. This may cause confusion in the interpretation of epigenetic marks for both features. The model plant Arabidopsis thaliana provides an excellent system to investigate composition and function of chromatin states at different levels of organization. In this review we will discuss recent developments in molecular networks underlying gene silencing and the relationship with visible heterochromatin in Arabidopsis.     

EMBRYO YELLOW gene, encoding a subunit of the conserved oligomeric Golgi complex, is required for appropriate cell expansion and meristem organization in Arabidopsis thaliana

We identified an embryo yellow ( eye ) mutation in Arabidopsis that leads to the abnormal coloration and morphology of embryos. The eye mutant formed bushy plants, with aberrant organization of the shoot apical meristem (SAM) and unexpanded leaves with irregular phyllotaxy. The epidermal cells of the eye mutant were much smaller than that of the wild-type. Thus, EYE is required for expansion of cells and organs, and for formation of the organized SAM. Hydrophobic layers of epidermal cells were also disrupted, suggesting that EYE might be involved in the generation of the extra-cellular matrix. The mutated gene encoded a protein that is homologous to Cog7, a subunit of the conserved oligomeric Golgi (COG) complex, which is required for the normal morphology and function of the Golgi appratus. The eye mutation caused mislocalization of a Golgi protein. In addition, the size of the Golgi apparatus was also altered. Thus, EYE might be involved in transport or retention of Golgi-localized proteins and in maintenance of Golgi morphology. We propose that some Golgi-localized proteins, distributions of which are controlled by EYE, play important roles in expansion of cells and organs, and in formation of the properly organized SAM in plants.     

Coexpression of neighboring genes in the genome of Arabidopsis thaliana

Large-scale analyses of expression data of eukaryotic organisms are now becoming increasingly routine. The data sets are revealing interesting and novel patterns of genomic organization, which provide insight both into molecular evolution and how structure and function of a genome interrelate. Our study investigates, for the first time, how genome organization affects expression of a gene in the Arabidopsis genome. The analyses show that neighboring genes are coexpressed. This pattern has been found for all eukaryotic genomes studied so far, but as yet, it remains unclear whether it is due to selective or nonselective influences. We have investigated reasons for coexpression of neighboring genes in Arabidopsis, and our evidence suggests that orientation of gene pairs plays a significant role, with potential sharing of regulatory elements in divergently transcribed genes. Using the data available in the KEGG database, we find evidence that genes in the same pathway are coexpressed, although this is not a major cause for the coexpression of neighboring genes.     

A subtilisin-like serine protease involved in the regulation of stomatal density and distribution in Arabidopsis thaliana

Stomata are specialized cellular structures in the epidermis of aerial plant organs that control gas exchange (H(2)O release and CO(2) uptake) between leaves and the atmosphere by modulating the aperture of a pore flanked by two guard cells. Stomata are nonrandomly distributed, and their density is controlled by endogenous and environmental factors. To gain insight into the molecular mechanisms regulating stomatal distribution, Arabidopsis thaliana mutants with altered stomatal characteristics were isolated and examined. The sdd1-1 mutant exhibits a two- to fourfold increase of stomatal density and formation of clustered stomata (i.e., stomata that are not separated by intervening pavement cells), whereas the internal leaf architecture is not altered. The SDD1 gene was identified by map-based cloning. It encodes a subtilisin-like serine protease related to prokaryotic and eukaryotic proteins. We propose that SDD1 acts as a processing protease involved in the mediation of a signal that controls the development of cell lineages that lead to guard cell formation.     

Instructive role of a peripheral pattern for the central patterning of the trigeminal projection at the brainstem and thalamus revealed by an artificially altered whisker pattern

The central patterning mechanism of neuronal circuits is an important issue in developmental neuroscience. We report here the role of a peripheral whisker pattern for the patterning of the trigeminal projection at the brainstem and thalamus in the mouse somatosensory system. The whisker pattern was manipulated by infecting the embryonic epidermis with adenovirus harboring Shh. The ectopic expression of Shh led to the induction of extra whiskers and displacement of whiskers, where these whiskers were histologically normal. The altered whisker pattern was isomorphically represented in the brainstem (barrelette: subnuclei principalis and subnuclei interpolaris), thalamus (barreloid) and cortex (barrel) as revealed by cytochrome oxidase staining. The barrelette-like pattern of the parvalbumin became discernible by immunostaining at P7 in subnuclei principalis and at P4 in subnuclei interpolaris in normal mice. These are the barrelette neurons projecting to the thalamus and the local circuit within the barrelette. The barrelette-like parvalbumin pattern also exhibits the altered whisker pattern induced by the adenovirus harboring Shh. These results highlight the role the peripheral whisker pattern for the central patterning of the brainstem, thalamus, and cortex in the mouse somatosensory system.     

Control of final seed and organ size by the DA1 gene family in Arabidopsis thaliana

Although the size of an organism is a defining feature, little is known about the mechanisms that set the final size of organs and whole organisms. Here we describe Arabidopsis DA1, encoding a predicted ubiquitin receptor, which sets final seed and organ size by restricting the period of cell proliferation. The mutant protein encoded by the da1-1 allele has a negative activity toward DA1 and a DA1-related (DAR) protein, and overexpression of a da1-1 cDNA dramatically increases seed and organ size of wild-type plants, identifying this small gene family as important regulators of seed and organ size in plants.     

Cloning and initial characterization of the Arabidopsis thaliana endoplasmic reticulum oxidoreductins

The oxidation and isomerization of disulfide bonds is necessary for the growth of all organisms. In yeast, the oxidative folding of secretory pathway proteins is catalyzed by protein disulfide isomerase (PDI), which requires Ero1p (endoplasmic reticulum oxidoreductin) for its own oxidation. In Homo sapiens, two homologues of Ero1p, Ero1-Lalpha and Ero1-Lbeta, have been cloned. Both Ero1-Lalpha and Ero1-Lbeta interact via disulfide bonds with PDI and support the oxidation of immunoglobulin light chains. However, the function of Ero proteins in plants has not yet been analyzed. In this article, we report the cloning of the two Ero1p homologues present in Arabidopsis thaliana, demonstrating that one of the cDNAs has a shorter terminal exon than predicted and differs from the annotated sequence found in the genome database. Sequence analysis of the Arabidopsis endoplasmic reticulum oxidoreductins (AEROs) reveals that both AERO1 and AERO2 are more closely related to each other than to either of the human Eros. Both in vitro translated AERO proteins are targeted to the endoplasmic reticulum and glycosylated. The ability to use a genetically tractable multicellular organism in combination with biochemical approaches should further our understanding of redox networks and Ero function in both plants and animals.     

A critical role of hypocretin deficiency in pregnancy

Hypocretin/orexin peptides are known for their role in the control of the wake–sleep cycle and narcolepsy–cataplexy pathophysiology. Recent studies suggested that hypocretin peptides also have a role in pregnancy. We tested this hypothesis by conducting a retrospective analysis on pregnancy complications in two different mouse models of hypocretin deficiency. We recorded 85 pregnancies of mice lacking either hypocretin peptides (knockout) or hypocretin‐releasing neurons (transgenic) and their wild‐type controls. Pregnancy was associated with unexplained dam death before delivery in 3/15 pregnancies in knockout mice, and in 3/23 pregnancies in transgenic mice. No casualties occurred in wild‐type pregnant dams (P < 0.007 versus hypocretin‐deficient mice as a whole). Hypocretin deficiency did not impact either on litter size or the number of weaned pups per litter. These data provide preliminary evidence of a critical role of hypocretin deficiency in pregnancy.