Origin and evolution of new exons in the rodent zinc finger protein 39 gene

The origin of new structures and functions is an important process in evolution. In the past decades, we have obtained some preliminary knowledge of the origin and evolution of new genes. However, as the basic unit of genes, the origin and evolution of exons remain unclear. Because young exons retain the footprints of origination, they can be good materials for studying origin and evolution of new exons. In this paper, we report two young exons in a zinc finger protein gene of rodents. Since they are unique sequences in mouse and rat genome and no homologous sequences were found in the orthologous genes of human and pig, the young exons might originate after the divergence of primates and rodents through exonization of intronic sequences. Strong positive selection was detected in the new exons between mouse and rat, suggesting that these exons have undergone significant functional divergence after the separation of the two species. On the other hand, population genetics data of mouse demonstrate that the new exons have been subject to functional constraint, indicating an important function of the new exons in mouse. Functional analyses suggest that these new exons encode a nuclear localization signal peptide, which may mediate new ways of nuclear protein transport. To our knowledge, this is the first example of the origin and evolution of young exons.

A zebrafish forebrain-specific zinc finger gene can induce ectopic dlx2 and dlx6 expression

Identifcation of the earliest forebrain-specific markers should facilitate the elucidation of molecular events underlying vertebrate forebrain determination and specification. Here we report the sequence and characterization of fez (forebrain embryonic zinc finger), a gene that is specifically expressed in the embryonic forebrain of zebrafish. Fez encodes a putative nuclear zinc finger protein that is highly conserved in Drosophila, zebrafish, Xenopus, mouse, and human. In zebrafish, the expression of fez becomes detectable at the anterior edge of the presumptive neuroectoderm by 70% epiboly. During the segmentation period, its expression is completely restricted to the rostral region of the prospective forebrain. At approximately 24 h postfertilization, fez expression is mostly confined to the telencephalon and the anterior-ventral region of the diencephalon. Although fez expression is present in one-eyed pinhead (oep) and cyclops (cyc) zebrfish mutants, the pattern is altered. Forced expression of fez induces ectopic expression of dlx2 and dlx6, two genes involved in brain development. Knockdown of fez function using a morpholino-based antisense oligo inhibited dlx2 expression in the ventral forebrain. Our studies indicate that fez is one of the earliest markers specific for the anterior neuroectoderm and it may play a role in forebrain development by regulating Dlx gene expression. (C) 2001 Academic Press.

A base-emitter self-aligned multi-finger Si1-xGex/Si power heterojunction bipolar transistor

With a crystal orientation dependent on the etch rate of Si in KOH-based solution, a base-emitter self-aligned large-area multi-linger configuration power SiGe heterojunction bipolar transistor (HBT) device (with an emitter area of about 880 mu m(2)) is fabricated with 2 mu m double-mesa technology. The maximum dc current gain is 226.1. The collector-emitter junction breakdown voltage BVCEO is 10 V and the collector-base junction breakdown voltage BVCBO is 16 V with collector doping concentration of 1 x 10(17) cm(-3) and thickness of 400 nm. The device exhibited a maximum oscillation frequency f(max) of 35.5 GHz and a cut-off frequency f(T) of 24.9 GHz at a dc bias point of I-C = 70 mA and the voltage between collector and emitter is V-CE = 3 V. Load pull measurements in class-A operation of the SiGe HBT are performed at 1.9 GHz with input power ranging from 0 dBm to 21 dBm. A maximum output power of 29.9 dBm (about 977 mW) is obtained at an input power of 18.5 dBm with a gain of 11.47 dB. Compared to a non-self-aligned SiGe HBT with the same heterostructure and process, f(max) and f(T) are improved by about 83.9% and 38.3%, respectively.

A Multi-Finger Si_(1-x)Ge_x/Si Heterojunction Bipolar Transistor for Wireless Power Amplifier Applications

A large area multi-finger configuration power SiGe HBT device(with an emitter area of about 880μm~2)was fabricated with 2μm double-mesa technology.The maximum DC current gain β is 214.The BV_(CEO) is up to 10V,and the BV_(CBO) is up to 16V with a collector doping concentration of 1×10~(17)cm~(-3) and collector thickness of 400nm.The device exhibits a maximum oscillation frequency f_(max) of 19.3GHz and a cut-off frequency f_T of 18.0GHz at a DC bias point of I_C=30mA and V_(CE)=3V.MSG(maximum stable gain)is 24.5dB,and U(Mason unilateral gain)is 26.6dB at 1GHz.Due to the novel distribution layout,no notable current gain fall-off or thermal effects are observed in the I-V characteristics at high collector current.

水稻小G蛋白Arf GTPase激活蛋白OsAGAP功能研究

　　锌指蛋白在植物生长发育中具有重要功能，它们可以识别并结合特定的DNA序列进行转录调控，还能够参与蛋白之间相互作用的调节。我们根据锌指蛋白等转录因子特征结构域的序列特点，从来自10 K水稻芯片的EST数据库中筛选出编码58个EST序列。通过对器官表达特异性的比较分析，从中选出七个只在单一器官表达的基因，并对这七个基因的功能进行研究。对其转基因水稻的表型分析发现，C1基因调节水稻的株高和穗的发育;LIM 家族的F9影响小花的形态，主要体现在雌蕊与雄蕊的发育;锌指蛋白S34调控叶倾角的变化;F14基因编码一个核定位的TFIIIA类锌指蛋白，具体功能尚不清楚;锌指蛋白F35转基因水稻主根缩短，侧根数目显著减少。它编码一个推测的ArfGAP (Arf GTPase activating protein)，据此我们将其命名为OsAGAP，并对其进行深入研究。 　　OsAGAP的cDNA全长为1328bp，编码的蛋白由320个氨基酸组成，含有两个保守结构域：锌指结构域和C2 结构域。其中锌指结构域属于CX2CX16CX2C类，即ArfGAP domain的特征结构。GTP酶活性测定试验表明，OsAGAP蛋白能够激活水稻Arf的GTP酶活性，另外，OsAGAP还能够恢复酵母ArfGAP缺失突变体的表型。说明OsAGAP编码的蛋白是水稻中的一个ArfGAP。 　　OsAGAP在水稻各器官中均有表达，但强弱有所不同。RNA原位杂交结果显示，它在茎尖分生组织与侧生原基及侧根部位表达强烈;它在根尖主要分布于中央维管组织、分生区、皮层细胞，最有趣的是恰好与生长素在根尖极性运输路径相吻合。在亚细胞水平，OsAGAP广泛分布于细胞膜、细胞质、细胞核。 　　OsAGAP超表达水稻主根、不定根长度缩短，侧根数目显著减少表现出类似于生长素极性运输突变体的表型。其主根伸长对TIBA的抑制作用不敏感，这暗示OsAGAP超表达水稻的生长素极性运输被破坏;另外，其对各种生长素的作用敏感性也发生变化，对IAA、2,4-D的不敏感，而对NAA的反应与野生型一致，根据各类生长素进出细胞机制不同，可以推测超表达水稻的输入能力存在缺陷。极性运输实验结果表明，超表达水稻极性运输能力被破坏;对生长素输入能力的测定进一步表明，超表达水稻根载体的介导的生长素输入能力显著下降。另外，NAA处理能够恢复超表达水稻中侧根发育受抑的表型缺陷。由此可见，OsAGAP在水稻中超表达破坏了生长素极性运输的输入能力。 　　FM1-43是一类特异标记囊泡运输的荧光染料。经其染色标记后，OsAGAP超表达水稻细胞内囊泡成片聚集，形成“BFA区间”，表现出囊泡运输被破坏的典型特征。透射电镜观察发现，超表达水稻细胞内有大量的小液泡，其中积累了电子密度很高的颗粒物质。由此推测，可能由于细胞的囊泡运输被破坏，导致胞内的代谢物质不能被正常运送或分泌，而在液泡中暂时贮存以维持细胞环境的稳定。 　　在酵母和动物细胞中的研究表明， ArfGAP是调控囊泡运输的一个重要因子，然而目前还没有关于ArfGAP在植物细胞中生理作用的报道。我们的结果说明，OsAGAP作为的一个ArfGAP，它通过调控水稻中的囊泡运输，而影响了生长素的极性运输，具体表现在对生长素输入能力的调控。由此，我们推测ArfGAP可能在生长素的极性运输中也起着重要的调控作用。 　　但OsAGAP在拟南芥中却通过调控植株生长素的水平，而影响了转基因拟南芥根的发育。每种生物都有多个ArfGAP，它们之间的分工存在联系，但各不相同。OsAGAP是拟南芥的外源基因，它在拟南芥中可能以不同于水稻的机制起作用。

OsLIC1基因参与水稻株型调控的功能研究

CCCH型锌指蛋白是进化上比较保守的一类锌指蛋白家族，其典型的氨基酸的基序为C-X7-8-C-X5-C-X3-H，其中X为任意氨基酸，这类锌指基序一般以重复的双拷贝形式存在。本论文克隆并鉴定了一个全新的、只含有一个CCCH型锌指基序的基因，利用反义RNA策略研究该基因功能，结果发现该基因的反义转基因植株表现出叶夹角增大的表型，因此我们将该基因命名为OsLIC1（Oryza sativa Lamina Increased Leaf Angle Control 1）。生物信息学分析发现该基因定位于水稻6号染色体近端粒的一端，位于BAC克隆AP004324中。OsLIC1与通常的CCCH型锌指蛋白含有多个重复的CCCH锌指基序不同，它只含有一个CCCH型锌指基序。除了CCCH锌指结构域以外，该蛋白在靠近C-端的位置还有一段丝氨酸（Ser）富集的区域，在此区域之前，还有一个在真核生物中相对保守的，以EELR为核心基序的结构域。采用基因枪将含OsLIC1-GFP融合构建的瞬时表达载体轰击入洋葱内表皮细胞，激光共聚焦显微镜观察发现OsLIC1-GFP可以定位到细胞核中。利用酵母转录激活系统发现以EELR为核心基序的结构域具有转录激活的功能。体外核酸结合活性分析显示OsLIC1蛋白可以结合双链DNA，这些结果证明OsLIC1是一个转录因子，这也是在植物中首次发现CCCH型锌指蛋白可以作为转录因子的方式调节基因的表达。 用玉米泛素启动子（Maize Ubiquitin promoter）驱动OsLIC1基因的反义表达载体转化水稻，获得的内源OsLIC1基因表达量下降的转基因植株表现出三个明显的表型：转基因植株的叶夹角增大;转基因的株高低于对照以及转基因植株的穗粒数减少。扫描电镜观察发现转基因植株叶夹角增大是由于近轴面细胞排列发生了改变以及维管束发育受阻引起的。转基因植株的Southern Blot和RT-PCR分析，结合Western Blot分析证明了转基因植株的表型与转基因事件之间的直接联系，并证明了转基因植株中内源OsLIC1在蛋白水平的确受到了抑制。采用RT-PCR技术、Promoter：：GUS和RNA in situ杂交三种方法相结合研究OsLIC1基因的表达模式，结果表明OsLIC1基因主要在叶颈、节以及分蘖原基中表达，这与转基因植株的表型相吻合，进一步证明了转基因植株的表型与基因功能之间的关系。Affymetrix 水稻全基因组芯片分析结果显示许多受油菜素内酯诱导表达的基因在转基因植株的叶颈材料中表达量上调，RT-PCR进一步验证了这一结果。由于在转基因植株中出现的叶夹角增大的表型和水稻油菜素内酯的作用相似，而基因芯片的结果又从分子水平提供了证据和线索。进一步采用RT-PCR和Promoter：：GUS相结合的方法研究OsLIC1基因对油菜素内酯的响应，结果发现OsLIC1基因可以被油菜素内酯诱导表达。而且，OsLIC1基因的反义转基因植株与野生型相比，表现出对油菜素内酯信号更敏感的响应。根据以上结果，推测OsLIC1可能是水稻油菜素内酯信号转导途径的负调控因子。水稻油菜素内酯合成和信号转导的突变体d2-1和d61-1具有直立的叶片的特征。用反义OsLIC1转基因植株以及野生型水稻与d2-1和d61-1突变体分别进行遗传杂交，结果发现在反义OsLIC1转基因植株与d2-1和d61-1突变体的杂交F1代中，都表现出叶夹角增大的表型。但是，在F2代中，在d2-1和d61-1纯合背景下，分别表现出叶夹角增大和叶片直立的表型，说明OsLIC1上位于d2-1，而d61-1则上位于OsLIC1。这一结果进一步证明了OsLIC1是通过参与水稻油菜素内酯信号调节而发挥对水稻叶夹角的调控作用。

水稻冷诱导基因OSCOIN的功能分析

对于双子叶模式植物拟南芥在逆境应答中的机理研究已取得了很大的进展，但在单子叶植物中的相关研究相对滞后。在单子叶植物水稻中仅仅报道了一些转录因子类基因以及与代谢有关的酶类基因，但与低温有关的分子伴侣、离子通道和载体等类基因的研究少见报道。 受低温诱导的水稻基因OsCOIN（AK104280）来自水稻10K cDNA芯片，它的cDNA全长有1593bp，开放阅读框内为1089bp，编码363个氨基酸，蛋白质的计算分子量42kDa，等电点5.25。在基因组序列中有7个外显子，6个内含子。生物信息分析显示，OsCOIN在第72—106氨基酸间形成一个指环结构域。OsCOIN蛋白没有跨膜区，定位于细胞质和细胞核。通过酵母双杂交实验证明了指环蛋白OsCOIN没有转录活性，故不是转录因子。RT-PCR结果表明，OsCOIN在所选的11种水稻组织中都有不同程度的表达，4C处理0.5h时OsCOIN基因开始较强地表达，持续较强地表达到4C处理48小时，72h时OsCOIN的表达量下降到起始的水平。另外,该基因表达还受ABA和盐诱导。 利用农杆菌介导的转化手段，得到三个OsCOIN超表达株系和六个RNAi株系。RNAi株系分蘖增多、植株矮化。为了分析OsCOIN与逆境胁迫的关系，实验中分析了超表达植株对低温等胁迫的耐性。结果表明，4C处理60h、72h、84h后，所有植株都出现萎蔫，恢复生长两周后超表达植株的存活率(分别为76.2%、71.4％和50％)明显高于野生型的存活率(分别为52.4%、22.2%和14.8%)。超表达OsCOIN水稻中，OsLti6b、OsNAC6和OsP5CS的表达量明显增加，而OsDhn1和OsDREB1a的表达量明显地降低，OsCDPK7和OsLti6a表达水平未受影响。 上述结果表明OsCOIN基因的过量表达抑制了OsDhn1和OsDREB1a的表达，促进了OsLti6b、OsNAC6和OsP5CS的表达。OsCOIN基因参与的低温响应途径与ABA相关，OsCOIN超表达植株不仅能耐低温，而且对盐和干旱胁迫有一定的抗性。

Xenopus RCOR2 (REST corepressor 2) interacts with ZMYND8, which is involved in neural differentiation

Regulation of neuronal gene expression is critical to nervous system development. REST (RE1-silencing transcription factor) regulates neuronal gene expression through interacting with a group of corepressor proteins including REST corepressors (RCOR). Here we show that Xenopus RCOR2 is predominantly expressed in the developing nervous system. Through a yeast two-hybrid screen, we isolated Xenopus ZMYND8 (Zinc finger and MYND domain containing 8) as an XRCOR2 interacting factor. XRCOR2 and XZMYND8 bind each other in co-immunoprecipitation assays and both of them can function as transcriptional repressors. XZMYND8 is co-expressed with XRCOR2 in the nervous system and overexpression of XZMYND8 inhibits neural differentiation in Xenopus embryos. These data reveal a RCOR2/ZMYND8 complex which might be involved in the regulation of neural differentiation. (C) 2010 Elsevier Inc. All rights reserved.

RNF220, an E3 ubiquitin ligase that targets Sin3B for ubiquitination

Modification of proteins by ubiquitination plays important roles in various cellular processes. During this process, the target specificity is determined by ubiquitin ligases. Here we identify RNF220 (RING finger protein 220) as a novel ubiquitin ligase for Sin3B. As a conserved RING protein, RNF220 can bind E2 and mediate auto-ubiquitination of itself. Through a yeast two-hybrid screen, we isolated Sin3B as one of its targets, which is a scaffold protein of the Sin3/HDAC (histone deacetylase) corepressor complex. RNF220 specifically interacts with Sin3B both in vitro and in vivo. Sin3B can be regulated by the ubiquitin-proteasome system. Co-expression of RNF220 promotes the ubiquitination and proteasomal degradation of Sin3B. Taken together, these results reveal a new mechanism for regulating the Sin3/HDAC complex. (C) 2010 Elsevier Inc. All rights reserved.

Sox and Zfx genes in giant panda

By using PCR cloning techniques, the DNA sequences of the HMG box regions of six Sox genes (pSox) and the zinc finger domains of two Zfx genes (pZfx) in the giant panda were identified. The giant panda Sox genes fell into two subfamilies, SOX-S1 and SOX-S2. The pSox and pZfx genes of the giant panda were highly homologous to the corresponding genes in mammals and revealed close substitution rates to those in the primates.

A new species of Amolops (Anura : Ranidae) from Southwest China

A new species of Amolops is described from a mountainous area of southern Yunnan Province, China. The species is unique in having a dark purple dorsum with small light yellow spots. The spots are smaller than the smallest finger disk. Other characters tha

Amolops bellulus: A new species of stream-breeding frog from western Yunnan, China (Anura : Ranidae)

We describe a new species of stream-breeding frog of the genus Amolops, Amolops bellulus, from the mountainous region of Lushui County, Yunnan Province, China, near the boundary between Yunnan and Myanmar. This species differs from all its congeners in a combination of morphological characteristics including the absence of a circummarginal groove at the tip of first finger, lack of a supratympanic fold, a white band along the upper jaw extending to shoulder, a black band starting from tip of snout along upper loreal region and extending to anterior flank of body, absence of a vocal sac, tympanum distinct and feebly concave, and the presence of vomerine teeth.