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PubMed2004

Comprehensive EST analysis of tomato and comparative genomics of fruit ripening.

A large tomato expressed sequence tag (EST) dataset (152 635 total) was analyzed to gain insights into differential gene expression among diverse plant tissues representing a range of developmental programs and biological responses. These ESTs were clustered and assembled to a total of 31 012 unique gene sequences. To better understand tomato gene expression at a plant system level and to identify differentially expressed and tissue-specific genes, we developed and implemented a digital expression analysis protocol. By clustering genes according to their relative abundance in the various EST libraries, expression patterns of genes across various tissues were generated and genes with similar patterns were grouped. In addition, tissues themselves were clustered for relatedness based on relative gene expression as a means of validating the integrity of the EST data as representative of relative gene expression. Arabidopsis and grape EST collections were also characterized to facilitate cross-species comparisons where possible. Tomato fruit digital expression data was specifically compared with publicly available grape EST data to gain insight into molecular manifestation of ripening processes across diverse taxa and resulted in identification of common transcription factors not previously associated with ripening.

Arabidopsis↗

PubMed2005

Osmotic regulation of root system architecture.

Although root system architecture is known to be highly plastic and strongly affected by environmental conditions, we have little understanding of the underlying mechanisms controlling root system development. Here we demonstrate that the formation of a lateral root from a lateral root primordium is repressed as water availability is reduced. This osmotic-responsive regulatory mechanism requires abscisic acid (ABA) and a newly identified gene, LRD2. Mutant analysis also revealed interactions of ABA and LRD2 with auxin signaling. Surprisingly, further examination revealed that both ABA and LRD2 control root system architecture even in the absence of osmotic stress. This suggests that the same molecules that mediate responses to environmental cues can also be regulators of intrinsic developmental programs in the root system.

Abscisic Acid↗

PubMed2006

Katanin's severing activity favors bundling of cortical microtubules in plants.

Higher plant cells exhibit interphase microtubule arrays specific to plants, which are essential for their developmental program. These cortical microtubules (CMT) consist of a population of highly dynamic microtubules that are usually organized into bundles in the cortex of the cells. The organization of CMT is intimately linked to the acquisition of specialized functions, and subsequentchanges in their distribution affect their properties. The mechanisms underlying the formation and the distribution of CMT are still unclear, and little is known about the proteins that are involved in this phenomenon. Here we investigated the putative role of katanin, the only known plant microtubule-severing protein, in the organization of CMT. We generated transgenic Arabidopsis lines that overexpress katanin under the control of an ethanol-inducible promoter. In response to an induced overexpression of katanin, CMT organized into numerous and thick bundles, which ultimately depolymerized. From the analyses of CMT patterns together with recent data on CMT dynamics, we propose that, in interphase cells, katanin's main activity is to free CMT, generating motile microtubules that incorporate into bundles.

Adenosine Triphosphatases↗

PubMed1980

Binary developmental commitments in normal and abnormal human morphogenesis.

An hypothesis is presented to account for the occurrence of overlapping patterns of anomalies in various malformation syndromes. According to this proposal, normal human morphogenesis occurs by means of a series of sequential commitments by neighboring groups of cells to alternative developmental programs.

Abnormalities, Multiple↗

PubMed1977

Analyses of stage-specific multiple forms of lactate dehydrogenase and of cytochrome c during spermatogenesis in the mouse.

Spermatogenesis is a programmed developmental process characterized by the inactivation of certain genes and the activation of other, testis-specific genes. Synthesis of unique gene products such as LDH-C4 and cytochrome ct occurs only at precise stage of germ cell formation. The developmental sequences of gene activation for these proteins was observed by immunohistochemical procedures. LDH-C4 is first detectable during mid-pachytene of the primary spermatocyte. The C subunits appear to be uniformly distributed throughout the cytoplasm of the spermatocyte. The mid-pachytene stage also marks the first appearance of cytochrome ct. The association of this electron transport protein with spermatocyte mitochondria is reflected in a granular fluorescence of specific antibody-treated sections of testis. Neither the C subunit of LDH, nor cytochrome ct appear in leptotene or early pachytene primary spermatocytes. These analyses indicate that there is stage-specific protein synthesis in the primary spermatocyte which is characterized by differential activation of the LDH-C locus and of the gene coding for cytochrome ct.

Age Factors↗

PubMed1985

Changes in the post-translational modification of lysosomal enzymes during development of Dictyostelium.

The form of post-translational modification present on two lysosomal enzymes--acid phosphatase and alpha-mannosidase--changes as part of the developmental program of Dictyostelium discoideum. Prior to 8 h of development, all enzyme molecules are of a single modification type (early form enzyme). Starting at 8 h of development, enzyme molecules with a second type of modification (late-form enzymes) begin to appear in the cell. We separated the early and late forms of these enzymes from each other by chromatography on DEAE-cellulose. We found that the change in protein modification affects the enzymes' in vitro properties. The early and late forms of both of these enzymes differ in thermostability and susceptibility to proteolytic inactivation. We also found that the late form of alpha-mannosidase is preferentially secreted. We suggest that by synthesizing molecules with a second form of modification, the cell confers new characteristics to its lysosomal enzymes.

Acid Phosphatase↗

PubMed1985

The regulation of 'early' enzymes during the development and dedifferentiation of Dictyostelium discoideum.

The specific activities of the enzymes alpha-mannosidase and N-acetylglucosaminidase increase immediately after the initiation of the development of bacterially grown cell cultures of Dictyostelium discoideum. The regulation of these two enzymes was found to be dissociable in the developmental timer mutant, FM-1, which aggregates 4.5 h earlier than wild-type cells due to the absence of the first rate-limiting component of the preaggregative period. The increase in alpha-mannosidase activity occurs in the absence of the first rate-limiting component, but the increase in N-acetylglucosaminidase activity does not. These results indicate the following: (1) the increase in the specific activity of alpha-mannosidase is not related to the timing of subsequent developmental stages; (2) the increase in the specific activity of N-acetylglucosaminidase is not necessary for the subsequent developmental program; and (3) either the increase in the specific activity of N-acetylglucosaminidase is dependent upon progress through the first rate-limiting component, or the increase in this enzyme activity and the first rate-limiting component are both dependent upon an early event for which FM-1 is defective. In addition to early development, we monitored the two enzyme activities during dedifferentiation. The results demonstrate that there is no difference between dedifferentiating wild-type cells and dedifferentiation-defective mutant HI-4 cells. Changes in enzyme specific activity accompanying dedifferentiation are dependent upon the composition of the dedifferentiation-inducing media and are consistent with the levels of these enzymes observed in cells growing in the different nutrient media.

Acetylglucosaminidase↗

PubMed1987

Expression of adult fast pattern of acetylcholinesterase molecular forms by mouse satellite cells in culture.

The pattern of acetylcholinesterase (AChE) molecular forms, obtained by sucrose gradient sedimentation, was studied at different in vitro developmental stages of myogenic cells isolated from adult mouse skeletal muscle. Only the globular forms were present in rapidly dividing satellite cells during the first days in culture. After myotube formation, a pattern similar to that described in mammalian fast-twitch skeletal muscle was observed. This pattern did not change during the following period in culture (up to 1 month) nor could it be modified by co-culturing with spinal cord motoneurons or by addition of brain-derived extracts. The internal-external localization of AChE molecular forms has been determined by the use of echothiophate iodide, a membrane-impermeant irreversible inhibitor of AChE. Echothiophate-treated cultures showed about 40% of both asymmetric and globular forms localized on the sarcolemma, with their active sites oriented outward. Analysis of culture medium from untreated cultures revealed the presence of both asymmetric and globular forms. When the same analysis was repeated on cultures of myoblasts derived from 16-day-old mouse embryos, the pattern of AChE forms was different. The myotubes derived from these cells exhibit a very small proportion of asymmetric form, which was not released into the medium. This pattern was not further modified during the following days of culture, nor by co-cultures with spinal cord motoneurons or by incubations with brain-derived extracts. Thus, the myotubes derived from myoblasts express in culture a clear phenotypic difference when compared to the corresponding myotubes from satellite cells, supporting the view that these two myogenic cells are endowed with different developmental programs.

Acetylcholinesterase↗

PubMed1989

Lamellar body formation precedes pulmonary surfactant apoprotein expression during embryonic mouse lung development in vivo and in vitro.

The purpose of this investigation was to determine whether lamellar inclusion body (LB) formation and surfactant apoprotein (SP-35) production are directly coordinated by temporal and positional information during development. In the present study we report a comparison between embryonic B10.A mouse lung morphogenesis and cytodifferentiation in vivo with that observed during organ culture in serumless medium. Precursor LB were first detected at embryonic day 12 (E12d), and progressively larger numbers and forms were produced during subsequent differentiation of respiratory alveolar duct epithelium. SP-35 was first detected during the canalicular period (E16.5d). Lung cultures (E12d) showed pseudoglandular and canalicular periods of morphogenesis, and both ciliated epithelial and type II cell differentiation. Nonciliated cells produced increasing numbers of lamellar inclusion bodies throughout the culture period. SP-35 was detected at 9 days in vitro (d.i.v.). These observations indicate (i) precursor LB formation precedes SP-35 expression and is not dependent on apoprotein synthesis; (ii) E12d lung development in vitro using serumless medium proceeds at a rate equivalent to 0.5 days in vivo through 11 d.i.v.; and (iii) morphogenesis and differentiation occur in the absence of exogenous hormones and growth factors. The cell-cell interactions that play a role in morphogenesis and cell differentiation appear to be intrinsic to the developmental program for embryonic lung development and are likely to be mediated by autocrine and/or paracrine factors.

Apoproteins↗

PubMed1990

Differential expression of muscle-specific enolase in embryonic and fetal myogenic cells during mouse development.

Three isoforms of the glycolytic enzyme enolase are present in mammals and birds. During development, a switch from the alpha to the beta form takes place in skeletal muscle. In order to investigate the molecular basis of this developmental transition of enolase isoforms, we extracted total RNA from limbs of mouse embryos of different ages, and from cultures of embryonic and fetal myogenic cells. The beta message was detected in limbs from 16-day-old fetuses by Northern-blot analysis and its level was found to increase in newborn and adult muscle; no significant amount of beta mRNA was present in samples from earlier developmental stages, which did however express high levels of the muscle-specific actin mRNA. Analysis of RNA extracted from embryonic and fetal myoblasts differentiated in culture revealed that the level of beta mRNA is about 9-fold higher in fetal myotubes than in embryonic myotubes, although the level of muscle actin is comparable in both types of myotubes. These results were confirmed by S1 nuclease protection experiments. Our data show that the appearance of beta enolase transcripts temporally correlates with the formation of the second generation of muscle fibers and suggest that the developmental transition from alpha to beta enolase is linked to a developmental program which takes place in fetal but not in embryonic muscle.

Animals↗

PubMed1991

Variable expression of retinoic acid receptor (RAR beta) mRNA in human oral and epidermal keratinocytes; relation to keratin 19 expression and keratinization potential.

Previous studies have revealed that the cells that form the different regions of the oral and epidermal stratified squamous epithelia represent a number of intrinsically distinct keratinocyte subtypes, each of which is developmentally programmed to preferentially express a particular pattern of keratins and type of suprabasal histology. Retinoic acid (RA) is known to modulate stratified squamous epithelial differentiation, including expression of the basal cell keratin K19 and the suprabasal keratins K1/K10 and K4/K13. We have found that all keratinocyte subtypes are similar in their steady state levels of RAR alpha and RAR gamma mRNAs in culture and that these levels are only minimally affected by RA. In contrast, RAR beta mRNA expression varies greatly among keratinocyte subtypes and, in eight of ten cell strains examined, directly correlated with their levels of K19 mRNA. Exposure to 10(-6) M RA increases the levels of RAR beta and K19 mRNA; conversely, complete removal of RA from the medium results in reduced levels of these messages. RA does not coordinately induce RAR beta and K19 messages in nonkeratinocyte cell types: fibroblasts cultured in the presence of 10(-6) M RA express very high levels of RAR beta mRNA but do not express detectable K19, and mesothelial cells decrease their levels of RAR beta and K19 mRNA in response to 10(-6) M RA. The correlation between RAR beta and K19 mRNA levels in most keratinocyte subtypes suggests a role for RAR beta in specifying patterns of keratin expression and suprabasal differentiation in stratified squamous epithelia.

Adult↗

PubMed1991

Developmental regulation of the inositol 1,4,5-trisphosphate phosphatases in Dictyostelium discoideum.

The cellular slime mold Dictyostelium discoideum is a microorganism in which growth and development are strictly separated. Starvation initiates a developmental program in which extracellular cAMP plays a major role as a signal molecule. In response to cAMP several second messengers are produced, including cAMP, cGMP and inositol 1,4,5-trisphosphate, (Ins(1,4,5)P3). Ins(1,4,5)P3 levels are controlled by the activation of phosphoinositidase C and the activity of the Ins(1,4,5)P3-degrading phosphatases. In Dictyostelium discoideum two major routes for the dephosphorylation of Ins(1,4,5)P3 are present: a 5-phosphatase, which hydrolyses Ins(1,4,5)P3 at the 5-position producing Ins(1,4)P2 as in vertebrate cells, and a 1-phosphatase which removes the 1-phosphate, giving Ins(4,5)P2, as in plants. In this paper we show that at the onset of development both the 1-phosphatase and the 5-phosphatase are present in equal amounts. During development the 5-phosphatase disappears leaving the 1-phosphatase as the single enzyme to remove Ins(1,4,5)P3. We conclude that during development Dictyostelium discoideum switches from a mixed type of Ins(1,4,5)P3 degradation to a more plant-like degradation pathway.

Chromatography, Ion Exchange↗

PubMed1991

Effect of the anti-microtubule drug oryzalin on growth and differentiation of the parasitic protozoan Leishmania mexicana.

The parasitic protozoan Leishmania mexicana differentiates from a non-motile intracellular amastigote in the mammalian macrophage phagolysosome into a motile, extracellular promastigote in the insect vector gut. This developmental program has been accomplished in vitro, thus providing a useful model for studying changes in the cytoskeleton during cell differentiation. The role of microtubules in leishmania differentiation was demonstrated by using the dinitroaniline herbicide oryzalin, which inhibited both leishmania proliferation and differentiation; 25 microM oryzalin reduced promastigote division by over 95%. Interestingly, at a sublethal dose (5 microM), promastigotes became round and multiflagellated but remained motile. At 50 microM oryzalin, the number of intracellular amastigotes decreased by 50%. However, leishmania differentiation seemed to be the most drug-sensitive stage: there was a 60% reduction in amastigote-to-promastigote differentiation at 0.5 microM oryzalin. The specific action of oryzalin on leishmania microtubules was verified by its inhibition of in vitro polymerization of leishmania microtubules, but not control mammalian microtubules (from rat brain). These findings indicate that microtubules play a major role in leishmania proliferation, maintenance of cell shape, and cytodifferentiation.

Animals↗

PubMed1992

Distribution of hyaluronan in the mouse endometrium during the periimplantation period of pregnancy.

The tissue distribution of stromal hyaluronan (HA) in the periimplantation mouse uterus was studied histochemically using a biotin-labelled HA-binding complex from cartilage proteoglycan. HA is present around proliferating stromal cells in both the pregnant and pseudopregnant mouse uterus prior to their differentiation into the decidualized phenotype. Decidualization is accompanied by clearance of HA from the extracellular matrix (ECM). This clearing is part of an intrinsic developmental program of the differentiating deciduum. A specific embryonic signal from the implanting conceptus is not required for this phenomenon to occur, since a similar response could be induced in deciduoma produced by artificial stimulation of a receptive uterus. Clearing of HA from the antimesometrial stroma is consistent with the hypothesis that the HA-negative decidual cell may be involved in restricting the invasion of trophoblast cells during embryo implantation. Retention of HA within angiogenic regions of the decidua basalis implies a functional role for this molecule in placental vascularization.

Animals↗

PubMed1992

Morphogenesis and differentiation of Dictyostelium cells interacting with immobilized glucosides: dependence on DIF production.

Previous work has shown that multicellular morphogenesis of submerged Dictyostelium cells is inhibited when they bind to glucosides covalently linked to polyacrylamide gels. The amoebae aggregate normally, but then the aggregates repeatedly disperse and reaggregate, whereas control cells go on to form tight aggregates. We have investigated the role of the stalk cell differentiation inducing factors (DIFs) in this process. In the presence of cyclic AMP, amoebae submerged at high cell density accumulate DIF and differentiate into stalk cells. We find that stalk cell differentiation is inhibited by interaction of the cells with glucoside gels in these conditions, but can be restored by the addition of exogenous DIF-1. Since the responsiveness of cells to DIF-1 is not altered, it appears likely that the effect of the glucoside gel is to block DIF-1 production. Further, the addition of DIF-1 or DIF-2 stimulates the formation of tight aggregates by cells developing on glucoside gels in the absence of cyclic AMP, thus preventing the rounds of aggregation and disaggregation otherwise seen. This suggests a role for DIF in morphogenesis as well as in controlling cell differentiation. We propose a model in which immobilized glucosides activate a specific receptor ("food sensor") which drives the amoebae toward the vegetative state and inhibits DIF accumulation. DIF, on the other hand, induces tight aggregate formation and so locks the amoebae into the developmental program.

Animals↗

PubMed1997

Purification, structure and in vitro molecular-chaperone activity of Artemia p26, a small heat-shock/alpha-crystallin protein.

Encysted brine-shrimp gastrulae bring their metabolism to a reversible standstill during diapause and quiescence, demonstrating a remarkable resistance to unfavourable environmental conditions. For example, mortality of Artemia embryos under normal temperature and hydration is very low, even after two years of anoxia, and embryos commonly experience complete desiccation as part of their developmental program. Previous evidence from our laboratories indicated that p26, an abundant low-molecular-mass cyst-specific protein capable of translocation into the nucleus, may have a protective function in Artemia cysts. p26 was purified to apparent homogeneity and a continuous sequence of 141 of its amino acids was determined by peptide sequencing, revealing that it is a member of the small-heat-shock/alpha-crystallin family of proteins. As determined by molecular-sieve chromatography and sucrose-density-gradient centrifugation, native p26 is a multimer of about 27 monomers with a molecular mass of approximately 700 kDa. Inactivation of citrate synthase was less when the enzyme was heated in the presence rather than the absence of p26. Additionally, the renaturation of heat-inactivated citrate synthase was promoted by p26. These results indicated that p26 possesses molecular-chaperone activity, a property of other small heat-shock/alpha-crystallin proteins. Our findings demonstrate that p26 has the potential to protect the macromolecular components of Artemia embryos, either as they encyst or upon exposure to environmental extremes. Protection may depend upon the ability of p26 to function as a molecular chaperone.

Amino Acid Sequence↗

PubMed2005

Functional specification in the Drosophila endoderm.

The discovery of homeobox gene clusters led us to realize that the mechanisms for body patterning and other developmental programs are evolutionally-conserved in vertebrates and invertebrates. The endoderm contributes to the lining of the gut and associated organs such as the liver and pancreas, which are critical for physiological functions. Our knowledge of endoderm development is limited; however, recent studies suggest that cooperation between the HNF3/Fork head and GATA transcription factors is crucial for endoderm specification. It is necessary to further understand the mechanism through which cells become functionally organized. Molecular genetic analyses of the Drosophila endoderm would provide insights into this issue. During proventriculus morphogenesis, a simple epithelial tube is folded into a functional multilayered structure, while two functions of midgut copper cells (i.e. copper absorption and acid secretion) can be easily visualized. The homeobox gene defective proventriculus (dve) plays key roles in these functional specifications.

Animals↗

PubMed1995

The role of the cytoskeleton in renal development.

The cytoskeleton is comprised of three separate filament networks: microtubules, microfilaments, and intermediate filaments. Collectively, these networks help establish and maintain the structural features of renal epithelial cells. During renal development, the cytoskeleton of the metanephric mesenchyme is extensively reorganized in order to create the cytoarchitectural elements that distinguish tubuloepithelial cells. This reorganization is coordinated with the formation of cell-cell contacts and cell-extracellular matrix interactions that are necessary to complete the developmental program. The actin cytoskeleton, microtubule network, and intermediate filament network all contribute to the development of polarity in the renal epithelial cells. The microtubule network determines the apical-basal axis of the cell. The actin cytoskeleton integrates topographic contacts between the cells and extracellular matrix. The tight junction and microvilli are subcellular structures that are associated with or comprised of actin filaments. Intermediate filament composition changes during the embryonic transition from metanephric mesenchyme to tubular epithelial cells. This review will describe the cell biology of the cytoskeletal elements in epithelial cells and the changes in cytoskeleton that accompany the formation of differentiated epithelial cells.

Actins↗