Single loci with major effects in fire ants : the effects of the complementary sex-determining locus and the green beard supergene on gene expression

Etant données la complexité et la redondance des réseaux de gènes influençant de nombreux phénotypes, l'étude des rares cas d'un locus unique ayant des effets importants sur de nombreux phénotypes peut fournir des informations cruciales sur l'évolution des traits complexes. Nous avons séquencé le génome de la fourmi de feu Solenopsis invicta pour étudier comment l'expression des gènes détermine les effets majeurs et étendus de deux loci uniques sur le phénotype. Le premier locus concerne la détermination du sexe par le modèle des allèles complémentaires. Ce locus est connu pour déterminer le sexe chez tous les hyménoptères mais n'a été caractérisé que chez les abeilles. Les hétérozygotes pour ce locus se développent en reines diploïdes (ou ouvrières stériles) alors que les homozygotes se développent en mâles diploïdes incapables de produire du sperme et les hémizygotes en mâles haploïdes fertiles. Nous avons comparé l'expression des gènes entre les reines et les deux types de mâles au stade pupe, ainsi que 1 et 11 jours après l'émergence. Nous avons trouvé un changement prononcé de l'expression des gènes chez les mâles diploïdes, passant de très proche de celle des reines au stade pupe à identique aux mâles haploïdes 11 jours après l'émergence. Cela signifie que les mâles diploïdes sont condamnés à être stériles parce que les effets après émergence du locus de détermination du sexe ne per¬mettent pas d'effacer les effets de la ploïdie sur l'expression des gènes pendant le stade pupe, quand la spermatogénèse prend place. Le second locus aux effets majeurs que nous avons étudié est le supergène dit "green beard", qui consiste en 616 gènes couvrant 55% d'un chromosome (13 Mb) et est caractérisé par une absence de recombinaison entre les deux variants du supergène : "Social B" et "Social b" (SB et Sb). Au travers de l'effet "green beard", par lequel les ouvrières avec le supergène Sb discriminent favorablement les reines qui partagent ce supergène de façon perceptible, le génotype des reines fondatrices au niveau de ce supergène détermine l'organisation de la colonie : soit elle contient une seule reine SB/SB, soit plusieurs reines SB/Sb. Nous avons montré que le chromosome Sb a évolué comme le chromosome Y, accumulant probablement des allèles favorables dans des colonies avec plusieurs reines mais défavorables dans des colonies avec une seule reine (cf. gènes sexuellement antagonistes), ainsi que des transposons et des séquences répéti¬tives. Nous avons également montré que le polymorphisme du supergène cause de grandes différences d'expression chez les ouvrières et particulièrement les reines mais pas chez les mâles. Pour comprendre comment le polymorphisme du supergène chez les reines peut affecter l'organisation de la colonie, nous avons comparé l'expression entre les génotypes SB/SB et SB/Sb chez des reines vierges (1 et 11 jours) et des reines matures. Nous avons montré que les reines SB/SB sur-régulent des gènes impliqués dans la reproduction, expli-quant pourquoi elle grandissent plus rapidement et peuvent fonder des colonies de façon indépendante, tandis que les reines SB/Sb (qui ne peuvent fonder une nouvelle colonie) sur-régulent des gènes de signalement chimique qui affectent l'organisation des colonies par l'effet "green beard". - Given the complexity and redundancy of the gene networks that underlie many pheno- types, the study of rare cases of a single locus having major effects on many phenotypes can give powerful insights into the evolution of complex traits. We sequenced the genome of Solenopsis invicta fire ants to study how gene expression mediates the widespread major effects of two single loci on phenotype. The first is the complementary sex-determining locus, which is known to exist in most Hymenoptera despite being characterized only for honeybees. Heterozygotes at this locus become diploid queens (or sterile workers), homozy¬gotes become aspermic diploid males, and hemizygotes become fertile haploid males. We compared gene expression between queens and both types of males in pupae and 1 and 11 days after eclosion. We found a pronounced shift in gene expression in diploid males, from being nearly identical to queens as pupae to identical to haploid males 11 days after eclosion. This means that diploid males are condemned to sterility because the overriding effects of the sex locus after eclosion cannot undo the ploidy effects on expression during the pupal stage, when spermatogenesis must be completed. The second locus with major ef¬fects that we studied was the so-called "green beard" supergene, which consists of 616 genes encompassing 55% of one chromosome (13 Mb), without recombination between the two variants "Social B" and "Social b" (SB and Sb) supergene. Through the green beard effect, i.e. workers with the Sb supergene discriminating in favor of queens who perceptibly share this supergene, the founding queen's genotype at the supergene determines colony organi¬zation: either headed by a single SB/SB queen or many SB/Sb queens. We show that the Sb chromosome evolved like a Y-chromosome, probably accumulating alleles beneficial in multi-queen colonies but disadvantageous in single-queen colonies (cf. sexually antagonistic genes), as well as transposons and repetitive sequences. We also show that the polymor¬phism of the supergene causes widespread expression differences in workers and especially queens but not in males. To understand how the polymorphism at the supergene in queen can transform colony organization, we compared the expression between SB/SB and SB/Sb virgin queens (1 and 11 days) and mother queens. We show that SB/SB queens up-regulate genes involved in reproduction, explaining why they mature faster and can found colonies independently, while SB/Sb queens (which cannot found colonies) up-regulate chemical signaling genes that can transform colonies through the green beard effect.

Perineuronal nets protect fast-spiking interneurons against oxidative stress.

A hallmark of schizophrenia pathophysiology is the dysfunction of cortical inhibitory GABA neurons expressing parvalbumin, which are essential for coordinating neuronal synchrony during various sensory and cognitive tasks. The high metabolic requirements of these fast-spiking cells may render them susceptible to redox dysregulation and oxidative stress. Using mice carrying a genetic redox imbalance, we demonstrate that extracellular perineuronal nets, which constitute a specialized polyanionic matrix enwrapping most of these interneurons as they mature, play a critical role in the protection against oxidative stress. These nets limit the effect of genetically impaired antioxidant systems and/or excessive reactive oxygen species produced by severe environmental insults. We observe an inverse relationship between the robustness of the perineuronal nets around parvalbumin cells and the degree of intracellular oxidative stress they display. Enzymatic degradation of the perineuronal nets renders mature parvalbumin cells and fast rhythmic neuronal synchrony more susceptible to oxidative stress. In parallel, parvalbumin cells enwrapped with mature perineuronal nets are better protected than immature parvalbumin cells surrounded by less-condensed perineuronal nets. Although the perineuronal nets act as a protective shield, they are also themselves sensitive to excess oxidative stress. The protection might therefore reflect a balance between the oxidative burden on perineuronal net degradation and the capacity of the system to maintain the nets. Abnormal perineuronal nets, as observed in the postmortem patient brain, may thus underlie the vulnerability and functional impairment of pivotal inhibitory circuits in schizophrenia.

The role of potassium in inflammasome activation by bacteria.

Many Gram-negative bacteria possess a type III secretion system (TTSS( paragraph sign)) that can activate the NLRC4 inflammasome, process caspase-1 and lead to secretion of mature IL-1beta. This is dependent on the presence of intracellular flagellin. Previous reports have suggested that this activation is independent of extracellular K(+) and not accompanied by leakage of K(+) from the cell, in contrast to activation of the NLRP3 inflammasome. However, non-flagellated strains of Pseudomonas aeruginosa are able to activate NLRC4, suggesting that formation of a pore in the cell membrane by the TTSS apparatus may be sufficient for inflammasome activation. Thus, we set out to determine if extracellular K(+) influenced P. aeruginosa inflammasome activation. We found that raising extracellular K(+) prevented TTSS NLRC4 activation by the non-flagellated P. aeruginosa strain PA103DeltaUDeltaT at concentrations above 90 mm, higher than those reported to inhibit NLRP3 activation. Infection was accompanied by efflux of K(+) from a minority of cells as determined using the K(+)-sensitive fluorophore PBFI, but no formation of a leaky pore. We obtained exactly the same results following infection with Salmonella typhimurium, previously described as independent of extracellular K(+). The inhibitory effect of raised extracellular K(+) on NLRC4 activation thus reflects a requirement for a decrease in intracellular K(+) for this inflammasome component as well as that described for NLRP3.

Dynamic regulation of notch 1 and notch 2 surface expression during T cell development and activation revealed by novel monoclonal antibodies.

It is well established that Notch signaling plays a critical role at multiple stages of T cell development and activation. However, detailed analysis of the cellular and molecular events associated with Notch signaling in T cells is hampered by the lack of reagents that can unambiguously measure cell surface Notch receptor expression. Using novel rat mAbs directed against the extracellular domains of Notch1 and Notch2, we find that Notch1 is already highly expressed on common lymphoid precursors in the bone marrow and remains at high levels during intrathymic maturation of CD4(-)CD8(-) thymocytes. Notch1 is progressively down-regulated at the CD4(+)CD8(+) and mature CD4(+) or CD8(+) thymic stages and is expressed at low levels on peripheral T cells. Immunofluorescence staining of thymus cryosections further revealed a localization of Notch1(+)CD25(-) cells adjacent to the thymus capsule. Notch1 was up-regulated on peripheral T cells following activation in vitro with anti-CD3 mAbs or infection in vivo with lymphocytic chorio-meningitis virus or Leishmania major. In contrast to Notch1, Notch2 was expressed at intermediate levels on common lymphoid precursors and CD117(+) early intrathymic subsets, but disappeared completely at subsequent stages of T cell development. However, transient up-regulation of Notch2 was also observed on peripheral T cells following anti-CD3 stimulation. Collectively our novel mAbs reveal a dynamic regulation of Notch1 and Notch2 surface expression during T cell development and activation. Furthermore they provide an important resource for future analysis of Notch receptors in various tissues including the hematopoietic system.

Involvement of connexin 43 in meiotic maturation of bovine oocytes.

In ovarian follicles, cumulus cells provide the oocyte with small molecules that permit growth and control maturation. These nutrients reach the germinal cell through gap junction channels, which are present between the cumulus cells and the oocyte, and between the cumulus cells. In this study the involvement of intercellular communication mediated by gap junction channels on oocyte maturation of in vitro cultured bovine cumulus-oocyte complexes (COCs) was investigated. The stages of oocyte maturation were determined by Hoechst 33342 staining, which showed that 90% of COCs placed in the maturation medium for 24 h progress to the metaphase II stage. Bovine COC gap junction communication was disrupted initially using n-alkanols, which inhibit any passage through gap junctions. In the presence of 1-heptanol (3 mmol l(-1)) or octanol (3.0 mmol l(-1) and 0.3 mmol l(-1)), only 29% of the COCs reached metaphase II. Removal of the uncoupling agent was associated with restoration of oocyte maturation, indicating that treatment with n-alkanols was neither cytotoxic nor irreversible. Concentrations of connexin 43 (Cx43), the major gap junction protein expressed in the COCs, were decreased specifically using a recombinant adenovirus expressing the antisense Cx43 cDNA (Ad-asCx43). The efficacy of adenoviral infection was > 95% in cumulus cells evaluated after infection with recombinant adenoviruses expressing the green fluorescence protein. RT-PCR performed on total RNA isolated from Ad-asCx43-infected COCs showed that the rat Cx43 cDNA was transcribed. Western blot analysis revealed a three-fold decrease in Cx43 expression in COCs expressing the antisense RNA for Cx43. Injection of cumulus cells with Lucifer yellow demonstrated further that the resulting lower amount of Cx43 in infected COCs is associated with a two-fold decrease in the extent of coupling between cumulus cells. In addition, oocyte maturation was decreased by 50% in the infected COC cultures. These results indicate that Cx43-mediated communication between cumulus cells plays a crucial role in maturation of bovine oocytes.

New Radicals and new applications of Dynamic Nuclear Polarization of biological systems

Dynamic Nuclear Polarization (DNP) is an emerging technique that could revolutionize the NMR study of small molecules at very low concentrations by the increase in sensitivity that results from transfer of polarization between electronic and nuclear spins. Although the underlying physics has been known for a long time, in the last few years there has been a lot of excitement on the chemistry and biology NMR community caused by the demonstration that the highly polarized nuclei that are prepared in solid state at very low temperatures (1-2 K) could be rapidly transferred to liquid samples at room temperature and studied in solution by conventional NMR techniques. In favorable cases several order of magnitude increases in sensitivity have been achieved. The technique is now mature enough that a commercial instrument is available. The efficiency of DNP depends on two crucial aspects: i) the efficiency of the nuclear polarization process and ii) the efficiency of the transfer from the initial solid state to the fluid state in which NMR is measured. The preferred areas of application (iii) will be dictated by situations in which the low concentration of the sample or its intrinsic low receptivity are the limiting factors .

Spermiogenesis particularities of a sperm storage species: Helicolenus dactylopterus (Teleostei: Scorpaenidae)

A study of the spermiogenesis and spermatozoa of Helicolenus dactylopterus was conducted. Females of this species have the capacity to store sperm within their ovaries, and male gametes have a considerable cytoplasmic mass surrounding their heads to survive the long period of intraovarian sperm storage. Our observations show that early spermatids are round-shaped cells and have a spherical nucleus with diffuse chromatin. The nuclear volume decreases as a result of progressive chromatin condensation during spermiogenesis, causing the nucleus to take on a U-shape. Flagellar insertion is not central to the nucleus but consistently occurs at an oblique angle towards one side of it. The flagellum is inserted into the nuclear fossa, without subsequent nuclear rotation. In mature spermatozoa, the flagellum is adjacent to the nucleus. A comparison of the spermatozoa in the testicular lobules and those in the intraovarian storage structures suggests that the increase in volume of the cytoplasmic mass may occur in the posterior region of the testis, in the testicular duct. Spermatozoa enter the ovary in groups that reach the ovarian lumen and are surrounded by the ovarian epithelium for storage in sperm storage crypts

Identification of Leishmania major cysteine proteinases as targets of the immune response in humans.

In this study, we report the identification of two parasite polypeptides recognized by human sera of patients infected with Leishmania major. Isolation and sequencing of the two genes encoding these polypeptides revealed that one of the genes is similar to the L. major cathepsin L-like gene family CPB, whereas the other gene codes for the L. major homologue of the cysteine proteinase a (CPA) of L. mexicana. By restriction enzyme digestion of genomic DNA, we show that the CPB gene is present in multiple copies in contrast to the cysteine proteinase CPA gene which could be unique. Specific antibodies directed against the mature regions of both types expressed in Escherichia coli were used to analyze the expression of these polypeptides in different stages of the parasite's life cycle. Polypeptides of 27 and 40 kDa in size, corresponding to CPA and CPB respectively, were detected at higher level in amastigotes than in stationary phase promastigotes. Purified recombinant CPs were also used to examine the presence of specific antibodies in sera from either recovered or active cases of cutaneous leishmaniasis patients. Unlike sera from healthy uninfected controls, all the sera reacted with recombinant CPA and CPB. This finding indicates that individuals having recovered from cutaneous leishmaniasis or with clinically apparent disease have humoral responses to cysteine proteinases demonstrating the importance of these proteinases as targets of the immune response and also their potential use for serodiagnosis.

Identification by a monoclonal antibody of a glycolipid highly expressed by cells from the human myeloid lineage.

A monoclonal antibody (MAb) HL-C5, which bound selectively to cells of the myeloid lineage tested, was derived from a fusion between P3/NS2/1-AG8 myeloma cells and splenocytes from a mouse immunized with cells of the promyelocytic leukemia line HL-60. Among a panel of 29 human cell lines derived from either hematopoietic or solid tumors, MAb HL-C5 was found to react exclusively with cells from the five differentiated acute myeloid leukemia lines, HL-60, ML1, ML2, ML3, KG-1B and not with the less differentiated myeloid lines. Fluorescence-activated cell sorter analysis of normal bone marrow samples confirmed that the reactivity of MAb HL-C5 was limited to myeloid cells, from the promyelocytic stage of differentiation to the mature granulocytes. Indirect immunoperoxidase staining of cytocentrifuge preparations of normal bone marrow and peripheral blood leukocytes confirmed these results and showed that MAb HL-C5 stained neutrophils but not eosinophils or basophils. The antigen recognized by HL-C5 was recovered in the upper phase of chloroform-methanol-water lipid extracts prepared from HL-60 cells. By competitive binding experiments, it was found that MAb HL-C5 recognizes the same antigenic determinant as MAb WGHS 29-1, which has been reported to react with glycolipids containing the sugar sequence lacto-N-fucopentaose 111. Autoradiographs of thin layer chromatograms of HL-60 glycolipid extracts which were revealed by incubation with MAb HL-C5 or WGHS 29-1 followed by the addition of 125I-labelled rabbit anti-mouse immunoglobulin antibody confirmed that the two MAbs reacted with the same or structurally very similar glycolipids.

Natural killer-like T cells develop in SJL mice despite genetically distinct defects in NK1.1 expression and in inducible interleukin-4 production.

An unusual subset of mature T cells expresses natural killer (NK) cell-related surface markers such as interleukin-2 receptor beta (IL-2R beta; CD122) and the polymorphic antigen NK1.1. These "NK-like" T cells are distinguished by their highly skewed V alpha and V beta repertoire and by their ability to rapidly produce large amounts of IL-4 upon T cell receptor (TCR) engagement. The inbred mouse strain SJL (which expresses NK1.1 on its NK cells) has recently been reported to lack NK1.1+ T cells and consequently to be deficient in IL-4 production upon TCR stimulation. We show here, however, that SJL mice have normal numbers of IL-2R beta+ T cells with a skewed V beta repertoire characteristic of "NK-like" T cells. Furthermore lack of NK1.1 expression on IL-2R beta+ T cells in SJL mice was found by backcross analysis to be controlled by a single recessive gene closely linked to the NKR-P1 complex on chromosome 6 (which encodes the NK1.1 antigen). Analysis of a panel of inbred mouse strains further demonstrated that lack of NK1.1 expression on IL-2R beta+ T cells segregated with NKR-P1 genotype (as assessed by restriction fragment length polymorphism) and thus was not restricted to the SJL strain. In contrast, defective TCR induced IL-4 production (which appeared to be a unique property of SJL mice) seems to be controlled by two recessive genes unlinked to NKR-P1. Collectively, our data indicate that "NK-like" T cells develop normally in SJL mice despite genetically distinct defects in NK1.1 expression and inducible IL-4 production.

A general definition and nomenclature for alternative splicing events

Understanding the molecular mechanisms responsible for the regulation of the transcriptome present in eukaryotic cells isone of the most challenging tasks in the postgenomic era. In this regard, alternative splicing (AS) is a key phenomenoncontributing to the production of different mature transcripts from the same primary RNA sequence. As a plethora ofdifferent transcript forms is available in databases, a first step to uncover the biology that drives AS is to identify thedifferent types of reflected splicing variation. In this work, we present a general definition of the AS event along with anotation system that involves the relative positions of the splice sites. This nomenclature univocally and dynamically assignsa specific ‘‘AS code’’ to every possible pattern of splicing variation. On the basis of this definition and the correspondingcodes, we have developed a computational tool (AStalavista) that automatically characterizes the complete landscape of ASevents in a given transcript annotation of a genome, thus providing a platform to investigate the transcriptome diversityacross genes, chromosomes, and species. Our analysis reveals that a substantial part—in human more than a quarter—ofthe observed splicing variations are ignored in common classification pipelines. We have used AStalavista to investigate andto compare the AS landscape of different reference annotation sets in human and in other metazoan species and found thatproportions of AS events change substantially depending on the annotation protocol, species-specific attributes, andcoding constraints acting on the transcripts. The AStalavista system therefore provides a general framework to conductspecific studies investigating the occurrence, impact, and regulation of AS.

A regulatory network for coordinated flower maturation.

For self-pollinating plants to reproduce, male and female organ development must be coordinated as flowers mature. The Arabidopsis transcription factors AUXIN RESPONSE FACTOR 6 (ARF6) and ARF8 regulate this complex process by promoting petal expansion, stamen filament elongation, anther dehiscence, and gynoecium maturation, thereby ensuring that pollen released from the anthers is deposited on the stigma of a receptive gynoecium. ARF6 and ARF8 induce jasmonate production, which in turn triggers expression of MYB21 and MYB24, encoding R2R3 MYB transcription factors that promote petal and stamen growth. To understand the dynamics of this flower maturation regulatory network, we have characterized morphological, chemical, and global gene expression phenotypes of arf, myb, and jasmonate pathway mutant flowers. We found that MYB21 and MYB24 promoted not only petal and stamen development but also gynoecium growth. As well as regulating reproductive competence, both the ARF and MYB factors promoted nectary development or function and volatile sesquiterpene production, which may attract insect pollinators and/or repel pathogens. Mutants lacking jasmonate synthesis or response had decreased MYB21 expression and stamen and petal growth at the stage when flowers normally open, but had increased MYB21 expression in petals of older flowers, resulting in renewed and persistent petal expansion at later stages. Both auxin response and jasmonate synthesis promoted positive feedbacks that may ensure rapid petal and stamen growth as flowers open. MYB21 also fed back negatively on expression of jasmonate biosynthesis pathway genes to decrease flower jasmonate level, which correlated with termination of growth after flowers have opened. These dynamic feedbacks may promote timely, coordinated, and transient growth of flower organs.

Extracellular matrix formation after transplantation of human embryonic stem cell-derived cardiomyocytes.

Transplantation of human embryonic stem cell-derived cardiomyocytes (hESC-CM) for cardiac regeneration is hampered by the formation of fibrotic tissue around the grafts, preventing electrophysiological coupling. Investigating this process, we found that: (1) beating hESC-CM in vitro are embedded in collagens, laminin and fibronectin, which they bind via appropriate integrins; (2) after transplantation into the mouse heart, hESC-CM continue to secrete collagen IV, XVIII and fibronectin; (3) integrin expression on hESC-CM largely matches the matrix type they encounter or secrete in vivo; (4) co-transplantation of hESC-derived endothelial cells and/or cardiac progenitors with hESC-CM results in the formation of functional capillaries; and (5) transplanted hESC-CM survive and mature in vivo for at least 24 weeks. These results form the basis of future developments aiming to reduce the adverse fibrotic reaction that currently complicates cell-based therapies for cardiac disease, and to provide an additional clue towards successful engraftment of cardiomyocytes by co-transplanting endothelial cells.

Evolutionary origin and functions of retrogene introns.

Retroposed genes (retrogenes) originate via the reverse transcription of mature messenger RNAs from parental source genes and are therefore usually devoid of introns. Here, we characterize a particular set of mammalian retrogenes that acquired introns upon their emergence and thus represent rare cases of intron gain in mammals. We find that although a few retrogenes evolved introns in their coding or 3' untranslated regions (untranslated region, UTR), most introns originated together with untranslated exons in the 5' flanking regions of the retrogene insertion site. They emerged either de novo or through fusions with 5' UTR exons of host genes into which the retrogenes inserted. Generally, retrogenes with introns display high transcription levels and show broader spatial expression patterns than other retrogenes. Our experimental expression analyses of individual intron-containing retrogenes show that 5' UTR introns may indeed promote higher expression levels, at least in part through encoded regulatory elements. By contrast, 3' UTR introns may lead to downregulation of expression levels via nonsense-mediated decay mechanisms. Notably, the majority of retrogenes with introns in their 5' flanks depend on distant, sometimes bidirectional CpG dinucleotide-enriched promoters for their expression that may be recruited from other genes in the genomic vicinity. We thus propose a scenario where the acquisition of new 5' exon-intron structures was directly linked to the recruitment of distant promoters by these retrogenes, a process potentially facilitated by the presence of proto-splice sites in the genomic vicinity of retrogene insertion sites. Thus, the primary role and selective benefit of new 5' introns (and UTR exons) was probably initially to span the often substantial distances to potent CpG promoters driving retrogene transcription. Later in evolution, these introns then obtained additional regulatory roles in fine tuning retrogene expression levels. Our study provides novel insights regarding mechanisms underlying the origin of new introns, the evolutionary relevance of intron gain, and the origin of new gene promoters.

Apoplastic polyesters in Arabidopsis surface tissues--a typical suberin and a particular cutin.

Cutinized and suberized cell walls form physiological important plant-environment interfaces as they act as barriers limiting water and nutrient loss and protect from radiation and invasion by pathogens. Due to the lack of protocols for the isolation and analysis of cutin and suberin in Arabidopsis, the model plant for molecular biology, mutants and transgenic plants with a defined altered cutin or suberin composition are unavailable, causing that structure and function of these apoplastic barriers are still poorly understood. Transmission electron microscopy (TEM) revealed that Arabidopsis leaf cuticle thickness ranges from only 22 nm in leaf blades to 45 nm on petioles, causing the difficulty in cuticular membrane isolation. We report the use of polysaccharide hydrolases to isolate Arabidopsis cuticular membranes, suitable for depolymerization and subsequent compositional analysis. Although cutin characteristic omega-hydroxy acids (7%) and mid-chain hydroxylated fatty acids (8%) were detected, the discovery of alpha,omega-diacids (40%) and 2-hydroxy acids (14%) as major depolymerization products reveals a so far novel monomer composition in Arabidopsis cutin, but with chemical analogy to root suberin. Histochemical and TEM analysis revealed that suberin depositions were localized to the cell walls in the endodermis of primary roots and the periderm of mature roots of Arabidopsis. Enzyme digested and solvent extracted root cell walls when subjected to suberin depolymerization conditions released omega-hydroxy acids (43%) and alpha,omega-diacids (24%) as major components together with carboxylic acids (9%), alcohols (6%) and 2-hydroxyacids (0.1%). This similarity to suberin of other species indicates that Arabidopsis roots can serve as a model for suberized tissue in general.