A recombinogenic targeting method to modify large-inserts for cis -regulatory analysis in transgenic mice: construction and expression of a 100-kb, zebrafish Hoxa-11b-lacZ reporter gene

Chiu, Chi Hua; Amemiya, Chris T.; Carr, Janet L.; Bhargava, Jaya; Hwang, Joseph; Shashikant, Cooduvalli S.; Ruddle, Frank H.; Wagner, Günter P.

doi:10.1007/s004270050016

Cited by 12 publications

(10 citation statements)

References 19 publications

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“…In this context it is interesting to note that we found an association between the presence of a putative repressor domain in the HOXA11 protein and the derived expression pattern (Chiu et al, 2000b). Similar clade specific differences in the noncoding regions of Hoxa-11 and Hoxa-13 are expected (Chiu et al, 2000a). (3) Some of the downstream target genes of Hoxa-11 and Hoxa-13 in tetrapods and fish are the same.…”

Section: Predictions and Implicationsmentioning

confidence: 85%

The tetrapod limb: A hypothesis on its origin

Wagner

Chiu

2001

J. Exp. Zool.

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A wrist joint and structures typical of the hand, such as digits, however, are absent in [Eustenopteron] (Andrews and Westoll, '68, p 240). Great changes must have been undergone during evolution of the ankle joint; the small number of large bones in the fin must somehow have developed into a large number of small bones, and it is very difficult to draw homologies in this region, or even be certain of what is being compared (Andrews and Westoll, '68, p 268). The tetrapod limb is one of the major morphological adaptations that facilitated the transition from an aquatic to a terrestrial lifestyle in vertebrate evolution. We review the paleontological evidence for the fin-limb transition and conclude that the innovation associated with evolution of the tetrapod limb is the zeugopodial-mesopodial transition, i.e., the evolution of the developmental mechanism that differentiates the distal parts of the limb (the autopodium, i.e., hand or foot) from the proximal parts. Based on a review of tetrapod limb and fish fin development, we propose a genetic hypothesis for the origin of the autopodium. In tetrapods the genes Hoxa-11 and Hoxa-13 have locally exclusive expression domains along the proximal-distal axis of the limb bud. The junction between the distal limit of Hoxa-11 expression and of the proximal limit of Hoxa-13 expression is involved in establishing the border between the zeugopodial and autopodial anlagen. In zebrafish, the expression domains of these genes are overlapping and there is no evidence for an autopodial equivalent in the fin skeleton. We propose that the evolution of the derived expression patterns of Hoxa-11 and Hoxa-13 may be causally involved in the origin of the tetrapod limb.

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Section: Predictions and Implicationsmentioning

confidence: 85%

The tetrapod limb: A hypothesis on its origin

Wagner

Chiu

2001

J. Exp. Zool.

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“…The pClasper homologous recombination technology has been used successfully in previous transgenic animal studies to investigate the evolution, regulation, and function of the mouse Hoxc8 gene (30)(31)(32), the zebrafish Hoxa-11b gene (33), and the mouse Dlx3 gene, among others (34,35). It proved essential for the construction of a functional Hec-6st-LacZ transgene capable of directing the tissue-specific expression of the LacZ reporter in register with endogenous Hec-6st gene expression.…”

Section: Discussionmentioning

confidence: 99%

Transgenic LacZ under control of Hec-6st regulatory sequences recapitulates endogenous gene expression on high endothelial venules

Liao

Bentley

Lebrun

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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Hec-6st is a highly specific high endothelial venule (HEV) gene that is crucial for regulating lymphocyte homing to lymph nodes (LN). The enzyme is also expressed in HEV-like vessels in tertiary lymphoid organs that form in chronic inflammation in autoimmunity, graft rejection, and microbial infection. Understanding the molecular nature of Hec-6st regulation is crucial for elucidating its function in development and disease. However, studies of HEV are limited because of the difficulties in isolating and maintaining the unique characteristics of these vessels in vitro. The novel pClasper yeast homologous recombination technique was used to isolate from a BAC clone a 60-kb DNA fragment that included the Hec-6st (Chst4) gene with flanking sequences. Transgenic mice were generated with the ␤-galactosidase (LacZ) reporter gene inserted in-frame in the exon II of Hec-6st within the isolated BAC DNA fragment. LacZ was expressed specifically on HEV in LN, as indicated by its colocalization with peripheral node vascular addressin. LacZ was increased in nasal-associated lymphoid tissue during development and was reduced in LN and nasal-associated lymphoid tissue by LT␤R-Ig (lymphotoxin-␤ receptor human Ig fusion protein) treatment in a manner identical to the endogenous gene.The transgene was expressed at high levels in lymphoid accumulations with characteristics of tertiary lymphoid organs in the salivary glands of aged mice. Thus, the Hec-6st-LacZ construct faithfully reproduces Hec-6st tissue-specific expression and can be used in further studies to drive expression of reporter or effector genes, which could visualize or inhibit HEV in autoimmunity.lymphoid organ ͉ pClasper ͉ peripheral node vascular addressin ͉ sulfotransferase ͉ lymphotoxin H igh endothelial venules (HEV) are the site of entry of naïve lymphocytes into the lymph node (LN) parenchyma from the blood stream. Lymphocyte tethering along HEV in LN is mediated by the interaction of L-selectin expressed on the surface of lymphocytes and L-selectin ligands expressed on HEV.

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“…All the colonies in 384-well microtiter plates were replica-plated onto nylon membranes for hybridization using standard techniques [49]. Two rounds of screening were performed.…”

Section: Methodsmentioning

confidence: 99%

Venom gland transcriptomes of two elapid snakes (Bungarus multicinctus and Naja atra) and evolution of toxin genes

et al. 2011

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BackgroundKraits (genus Bungarus) and cobras (genus Naja) are two representative toxic genera of elapids in the old world. Although they are closely related genera and both of their venoms are very toxic, the compositions of their venoms are very different. To unveil their detailed venoms and their evolutionary patterns, we constructed venom gland cDNA libraries and genomic bacterial artificial chromosome (BAC) libraries for Bungarus multicinctus and Naja atra, respectively. We sequenced about 1500 cDNA clones for each of the venom cDNA libraries and screened BAC libraries of the two snakes by blot analysis using four kinds of toxin probes; i.e., three-finger toxin (3FTx), phospholipase A2 (PLA2), kunitz-type protease inhibitor (Kunitz), and natriuretic peptide (NP).ResultsIn total, 1092 valid expressed sequences tags (ESTs) for B. multicinctus and 1166 ESTs for N. atra were generated. About 70% of these ESTs can be annotated as snake toxin transcripts. 3FTx (64.5%) and β bungarotoxin (25.1%) comprise the main toxin classes in B. multicinctus, while 3FTx (95.8%) is the dominant toxin in N. atra. We also observed several less abundant venom families in B. multicinctus and N. atra, such as PLA2, C-type lectins, and Kunitz. Peculiarly a cluster of NP precursors with tandem NPs was detected in B. multicinctus. A total of 71 positive toxin BAC clones in B. multicinctus and N. atra were identified using four kinds of toxin probes (3FTx, PLA2, Kunitz, and NP), among which 39 3FTx-postive BACs were sequenced to reveal gene structures of 3FTx toxin genes.ConclusionsBased on the toxin ESTs and 3FTx gene sequences, the major components of B. multicinctus venom transcriptome are neurotoxins, including long chain alpha neurotoxins (α-ntx) and the recently originated β bungarotoxin, whereas the N. atra venom transcriptome mainly contains 3FTxs with cytotoxicity and neurotoxicity (short chain α-ntx). The data also revealed that tandem duplications contributed the most to the expansion of toxin multigene families. Analysis of nonsynonymous to synonymous nucleotide substitution rate ratios (dN/dS) indicates that not only multigene toxin families but also other less abundant toxins might have been under rapid diversifying evolution.

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A recombinogenic targeting method to modify large-inserts for cis -regulatory analysis in transgenic mice: construction and expression of a 100-kb, zebrafish Hoxa-11b-lacZ reporter gene

Cited by 12 publications

References 19 publications

The tetrapod limb: A hypothesis on its origin

The tetrapod limb: A hypothesis on its origin

Transgenic LacZ under control of Hec-6st regulatory sequences recapitulates endogenous gene expression on high endothelial venules

Venom gland transcriptomes of two elapid snakes (Bungarus multicinctus and Naja atra) and evolution of toxin genes

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