The Hox Paradox: More complex(es) than imagined

An understanding of the origin of different body plans requires knowledge of how the genes and genetic pathways that control embryonic development have evolved. The Hox genes provide an appealing starting point for such studies because they play a well-understood causal role in the regionalization of the body plan of all bilaterally symmetric animals. Vertebrate evolution has been characterized by gene, and possibly genome, duplication events, which are believed to have provided raw genetic material for selection to act upon. It has recently been established that the Hox gene organization of ray-finned fishes, such as the zebrafish, differs dramatically from that of their lobe-finned relatives, a group that includes humans and all the other widely used vertebrate model systems. This unusual Hox gene organization of zebrafish is the result of a duplication event within the ray-finned fish lineage. Thus, teleosts, such as zebrafish, have more Hox genes arrayed over more clusters (or "complexes") than do tetrapod vertebrates. Here, I review our understanding of Hox cluster architecture in different vertebrates and consider the implications of gene duplication for Hox gene regulation and function and the evolution of different body plans.     

Retinoic acid affects patterning along the anterior–posterior axis of the ascidian embryo

Because retinoic acid (RA) is known to affect anterior-posterior patterning in vertebrate embryos, it was questioned whether it shows similar effects in a more primitive chordate, the ascidian Halocynthia roretzi . Ascidian embryos treated with RA exhibited truncated phenotypes in a dose-dependent manner similar to the anterior truncations seen in vertebrate embryos. The most severely affected larvae possessed a round trunk without the papillae characteristic of the anterior terminal epidermis. Retinoic acid also altered the expression of HrHox-1 and Hroth in a dose-dependent manner. Expression of HrHox-1 increased, whereas expression of Hroth decreased with increasing levels of RA. In treated embryos, HrHox-1 was first expressed pan-ectodermally, then degraded in all but specific regions of the embryo. By contrast, initiation of Hroth expression was not affected, but epidermal expression was lost while expression in the neural tube narrowed toward the anterior in tail-bud embryos. These alterations in the expression of homeobox genes appear to correlate closely to the morphological defects elicited by RA treatment, suggesting broad conservation of developmental patterning mechanisms within the Phylum Chordata.     

The fates of zebrafish Hox gene duplicates

In his 1970 book, Susumu Ohno stressed the importance of gene duplication in the evolution of the vertebrate genome and body plan. He elaborated the idea that duplication events provide novel genetic material on which evolution may act. Data are accumulating to show that extensive duplication events, perhaps incorporating the duplication of entire genomes, occurred in the lineage leading to teleost fishes. These duplications may have been pivotal in the explosive radiation of this highly successful vertebrate group. Thus, the teleosts provide us with an ideal opportunity to investigate the fates and functions of duplicated genes. A convenient system for these studies is the zebrafish, Danio rerio, which has become a popular genetic and embryological model.     

Vertebrate hox temporal collinearity: does it exist and what is it’s function?

Hox temporal collinearity (TC) is a mysterious feature of embryogenesis. This article is opportune because of a recent challenge to TC’s existence This challenge is examined and the evidence that TC does exist is presented. Its function is discussed. Temporal collinearity is thought to be important because it lays the basis for Hox spatial collinearity and the vertebrate A-P axial pattern. The time-space translation mechanism whereby this occurs is examined.     

All-trans-retinoic Acid Reduces BACE1 Expression under Inflammatory Conditions via Modulation of Nuclear Factor κB (NFκB) Signaling

Insulin resistance and neuroinflammation have emerged as two likely key contributors in the pathogenesis of Alzheimer disease (AD), especially in those sporadic AD cases compromised by diabetes or cardiovascular disease. Amyloid-β (Aβ) deposition and its associated inflammatory response are hallmarks in sporadic AD brains. Elevated expression and activity of β-secretase 1 (BACE1), the rate-limiting enzyme responsible for the β-cleavage of amyloid precursor proteins to Aβ peptides, are also observed in sporadic AD brains. Previous studies have suggested that there is therapeutic potential for retinoic acid in treating neurodegeneration based on decreased Aβ. Here we discovered that BACE1 expression is elevated in the brains of both Tg2576 transgenic mice and mice on high fat diets. These conditions are associated with a neuroinflammatory response. We found that administration of all-trans-retinoic acid (atRA) down-regulated the expression of BACE1 in the brains of Tg2576 mice and in mice fed a high fat diet. Moreover, in LPS-treated mice and cultured neurons, BACE1 expression was repressed by the addition of atRA, correlating with the anti-inflammatory efficacy of atRA. Mutations of the NFκB binding site in BACE1 promoter abolished the suppressive effect of atRA. Furthermore, atRA disrupted LPS-induced nuclear translocation of NFκB and its binding to BACE1 promoter as well as promoting the recruitment of the corepressor NCoR. Our findings indicate that atRA represses BACE1 gene expression under inflammatory conditions via the modulation of NFκB signaling.     

Glucocorticoid and retinoid regulation of alpha-2 type I procollagen promoter activity.

Glucocorticoids decrease type I procollagen synthesis by decreasing the steady state levels of procollagen mRNAs and mRNA synthesis. The present studies were undertaken to determine the functional sequences of the pro alpha 2(I) collagen gene required for the glucocorticoid-mediated decrease of type I procollagen mRNA synthesis. Embryonic mouse fibroblasts were stably transfected with the pR40 DNA CAT construct containing the 5' flanking region fragment from -2048 to +54 and the intronic fragment from +418 to +1524 of the mouse alpha 2(I) collagen gene. Dexamethasone treatment of these pR40 transfected fibroblasts resulted in a significant decrease in CAT activity which agrees with the glucocorticoid-mediated decrease of the steady state levels of type I procollagen mRNAs. To determine the possible role of the first intron fragment in the dexamethasone-mediated decrease of CAT activity, pR36, a CAT plasmid containing the first intron fragment and the SV40 early promoter, was transfected into mouse fibroblasts and treated with dexamethasone. No significant decrease in CAT activity was observed. The dexamethasone-mediated response was then localized within the 5' flanking region by preparing a series of constructs containing internal deletions and transfecting these plasmids into mouse fibroblasts. The regions -2048 to -981 and -506 to -351 were required for the dexamethasone response of gene activity. However, the DNA stretch from -981 to -506 was not. Analysis of the DNA sequences of these regions revealed a single GRE at -1023 to -1018 and a modified doublet at -873 to -856.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of retinoic acids on the dendritic morphology of cultured hippocampal neurons

Vitamin A-derived retinoic acids (RAs) are known to exert a variety of biological actions, including modulatory effects on cell differentiation and apoptosis. A recent study has demonstrated that 13- cis -RA and all- trans -RA suppressed neurogenesis in the dentate gyrus of the hippocampus in adult mice. The present experiments were performed to see whether 13- cis -RA and all- trans -RA could alter the dendritic morphology of cultured hippocampal neurons via RA receptors: retinoic acid receptor (RAR) and retinoid X receptor (RXR). High doses of 13- cis -RA and all- trans -RA exerted a negative effect on the cultured hippocampal neurons, while a low dose of 13- cis -RA but not all- trans -RA caused a positive effect. The negative changes induced by 13- cis -RA and all- trans -RA were antagonized by RXR antagonists and RAR antagonists, respectively. The positive changes induced by a low dose of 13- cis -RA were blocked by both RXR antagonists and RAR antagonists. These results suggest that RAs at high concentrations cause a negative effect on the dendritic morphology of cultured hippocampal neurons through RA receptors, while RAs at low concentrations exert a positive influence on cultured hippocampal neurons.     

Comparison of Models for the Collinearity of Hox Genes in the Developmental Axes of Vertebrates

Hox gene clusters are very frequent in many animal genomes and their role in development is pivotal. Particularly in vertebrates, intensive efforts have established several properties of Hox clusters. The collinearity of Hox gene expressions (spatial, temporal and quantitative) is a common feature of the vertebrates. During the last decade, genetic engineering experiments have revealed some important facets of collinearity during limb and trunk development in mice. Two models have been proposed to explain all these properties. On one hand the ‘two-phases model’ makes use of the molecular regulatory mechanisms acting on the Hox genes. On the other hand, the’biophysical model’ is based on the signals transduced inside the cell nucleus and the generation of forces which apply on the cluster and lead to a coordinated activation of Hox genes. The two models differ fundamentally and a critical and detailed comparison is presented. Furthermore, experiments are proposed for which the two models provide divergent predictions. The outcome of these experiments will help to decide which of the two models is valid (if any).     

Retinoid dynamics in chicken eye during pre-and postnatal development

Changes in the steady state level of retinols, retinaldehydes and retinyl esters in the trans and 11-cis forms and trans retinoic acid were measured in whole chicken eye during development from day 6in ovo to day 3 post-hatch. These retinoids, quantified by different HPLC systems, were detected in this time sequence: trans-retinol and trans-retinyl esters in the first weekin ovo, 11-cis-retinol in the second week. The highest level of 11-cis-retinaldehyde and 11-cis-retinyl esters was reached at the end of developmentin ovo; however, their levels increased further after hatching. The retinoic acid level decreased at the end of the first week, rising again at the end of the second week.The enzyme activities involved in the metabolism of these retinoids-acyl-CoA: retinol acyltransferase, trans-retinol dehydrogenase, 11-cis-retinol dehydrogenase, trans-retinyl ester hydrolase and trans: 11-cis-retinol isomerase were also estimated and they were detectable already in the first week of developmentin ovo.At day 6 of the biosynthesis of retinoic acid by the retinaldehyde dehydrogenase activity from retina cytosol was also shown.     

Hox gene expression reveals regionalization along the anteroposterior axis of the zebrafish notochord

 The vertebrate Hox genes have been shown to confer regional identity along the anteroposterior axis of the developing embryo, especially within the central nervous system (CNS) and the paraxial mesoderm. The notochord has been shown to play vital roles in patterning adjacent tissues along both the dorsoventral and mediolateral axes. However, the notochord’s role in imparting anteroposterior information to adjacent structures is less well understood, especially as the notochord shows no morphological distinctions along the anteroposterior axis and is not generally described as a segmental or compartmentalized structure. Here we report that four zebrafish hox genes: hoxb1, hoxb5, hoxc6 and hoxc8 are regionally expressed along the anteroposterior extent of the developing notochord. Notochord expression for each gene is transient, but maintains a definite, gene-specific anterior limit throughout its duration. The hox gene expression in the zebrafish notochord is spatially colinear with those genes lying most 3’ in the hox clusters having the most anterior limits. The expression patterns of these hox cluster genes in the zebrafish are the most direct molecular evidence for a system of anteroposterior regionalization of the notochord in any vertebrate studied to date. Received: 30 March 1998 / Accepted: 16 June 1998     

Retinoic Acid Improves a Hybridoma Culture in a Fructose-Based Medium by Up-Regulation of Fructose Incorporation via Retinoid Nuclear Receptors

Fructose was focused on as an alternative sugar source to glucose in a hybridoma culture medium because it decreases lactate production during cultivation, leading to cell and product stability. But, not all human hybridoma cell lines grew well in a fructose-based serum-free medium. We found that the addition of all-trans-retinoic acid to the fructose-based medium improved the growth and monoclonal antibody production of hybridoma cell lines by up-regulation of fructose incorporation that represented increased expression of the fructose transporter, GLUT5. Selective activation of retinoid nuclear receptor by synthetic ligands showed that both retinoic acid receptors and retinoid X receptors might be related to the improvement of the fructose-based hybridoma culture. This study might be applicable to cell cultures susceptible to lactate and pH changes as well as hybridoma cultures.     

qPCR array检测分析 C57BL/6小鼠胚胎后肢发育基因表达谱

目的:胚胎生育过程中因肢体发育异常造成的出生缺陷比率不低,其相关基因表达模式尚不明确。本实验通过建立实时定量PCR芯片(Real-time quantitative polymerasechain reaction array,qPCR array)检测方案,研究C57BL/6品系小鼠后肢发育相关基因的表达谱。方法:以同源异形盒基因家族(Hox)、Wnt5a、配对同源结构域基因(Pitx1)、成纤维生长因子(Fgf8)、音猬因子(Shh)等小鼠肢体发育相关的重要基因制作基因检测表达谱,以C57BL/6品系怀孕雌鼠为材料,取胚胎肢芽发育的四个关键时期(E10.5,E11.5,E12.5,E13.5)的胎鼠后肢,利用qPCR array方案检测表达谱中基因的相对表达水平差异。结果:通过已建立的qPCR array检测了C57BL/6品系小鼠胚胎后肢发育时期Hox家族、Wnt5a、Pitx1、Fgf8、Shh等基因的表达差异。以E10.5为对照,检测出在后肢发育时期基因呈三种表达模式,即Hoxb6、Hoxb8、Hoxc8、Hoxc9、Hoxc10、Hoxd9和Shh基因的表达水平呈上调;Hoxa11、Hoxa13、Hoxc12、Hoxc13、Hoxd13等基因表达出现下调;Hoxc9、Hoxc10、Hoxc11、Hoxd9、Hoxd12、Fgf8和Pitx1等基因的相对表达量呈先上调后下调的曲线表达模式,且有少部分基因在小鼠后肢发育时期表达水平无明显变化。结论:Hox家族、Wnt5a、Pitx1、Fgf8、Shh等基因在小鼠后肢发育时期表达,并且表达模式存在明显差异。     

Vitamin A deficiency results in the dose-dependent acquisition of anterior character and shortening of the caudal hindbrain of the rat embryo

The developing nervous system is particularly vulnerable to vitamin A deficiency. Retinoid has been proposed to be a posteriorizing factor during hindbrain development, although direct evidence in the mammalian embryo is lacking. In the present study, pregnant vitamin A-deficient (VAD) rats were fed purified diets containing varying levels of all-trans-retinoic acid (atRA; 0, 0.5, 1.5, 6, 12, 25, 50, 125, or 250 microg/g diet) or were supplemented with retinol. Hindbrain development was studied from embryonic day 10 to 12.5 ( approximately 6 to 40 somites). Normal morphogenesis was observed in all embryos from groups fed 250 microg atRA/g diet or retinol. The most caudal region of the hindbrain was the most sensitive to retinoid insufficiency, as evidenced by a loss of the hypoglossal nerve (cranial nerve XII) in embryos from the 125 microg atRA/g diet group. Further reduction of atRA to 50 microg/g diet led to the loss of cranial nerves IX, X, XI, and XII and associated sensory ganglia IX and X in all embryos as well as the loss of hindbrain segmentation caudal to the rhombomere (r) 3/4 border in a subset of embryos. Dysmorphic orthotopic otic vesicles or immature otic-like vesicles in both orthotopic and caudally ectopic locations were also observed. As the level of atRA was reduced, a loss of caudal hindbrain segmentation was observed in all embryos and the incidence of otic vesicle abnormalities increased. Perturbations in hindbrain segmentation, cranial nerve formation, and otic vesicle development were associated with abnormal patterning of the posterior hindbrain. Embryos from VAD dams fed between 0.5 and 50 microg atRA/g diet exhibited Hoxb-1 protein expression along the entire neural tube caudal to the r3/r4 border at a time when it should be restricted to r4. Krox-20 protein expression was expanded in r3 but absent or reduced in presumptive r5. Hoxd-4 mRNA expression was absent in the posterior hindbrain, and the rostral limit of Hoxb-5 protein expression in the neural tube was anteriorized, suggesting that the most posterior hindbrain region (r7/r8) had been deleted and/or improperly patterned. Thus, when limiting amounts of atRA are provided to VAD dams, the caudal portion of the hindbrain is shortened and possesses r4/r5-like characteristics, with this region finally exhibiting r4-like gene expression when retinoid is restricted even more severely. Thus, regions of the anterior hindbrain (i.e., r3 and r4) appear to be greatly expanded, whereas the posterior hindbrain (r5-r8) is reduced or absent. This work shows that retinoid plays a critical role in patterning, segmentation, and neurogenesis of the caudal hindbrain and serves as an essential posteriorizing signal for this region of the central nervous system in the mammal.