Tyrosine Aminotransferase Gene (SmTAT) Revealed Genetic Diversity and Phylogeny of Cultivated Danshen (Salvia miltiorrhiza) Populations

Chinese traditional medicine Danshen is the radix of the perennial herbs of Salvia miltiorrhiza Bunge, which has a variety of pharmacological effects and is traditionally and extensively applied clinically to treat cardiovascular disorders. In this research, the genomic genes for tyrosine aminotransferase (TAT) of 38 cultivated populations of Danshen in China were cloned and bioinformatic analyses were conducted to reveal its genetic diversity and phylogeny. The full-length SmTAT was 2296 - 2444 bp including 6 exons (encoding 411 amino acids) and 5 introns. Overall, the SmTAT genes in cultivated Danshen populations are highly conserved with a relative low level of genetic diversity. The spliced exons (1236 bp) had 23 SNP variations with a rate of 1.86%, of which 22 occurred in the white flower S. miltiorrhiza Bge.f.alba population (W-SCHY-W-1) and led to 5 amino acid variations. The entire 290 SNP variations with a rate of 24% in the 5 introns occurred exclusively in W-SCHY-W-1. Phylogenetic trees based on the full-length, combined introns, the spliced exons, and the deduced amino acid sequences of SmTAT all showed a two-clade basic structure with W-SCHY-W-1 uniquely standing alone. The SmTAT gene of the white flower population (W-SCHY-W-1) is unique and especially rich in variations. The first time clarified genomic SmTAT gene structure and genetic diversity in cultivated Danshen populations laid an excellent foundation for further studies on the biosynthesis of bioactives and the molecular breeding of Danshen as well as in plant tyrosine metabolism.

26S proteasome inhibitors inhibit dexamethasone-dependent increase of tyrosine aminotransferase and tryptophan 2,3-dioxygenase mRNA levels in primary cultured rat hepatocytes

Dexamethasone (Dex), a ligand for transcriptional enhancement of tyrosine aminotransferase (TAT) and tryptophan 2,3-dioxygenase (TO) genes, (100 nM) maximally increased these mRNA levels at 12 h and 7 h in primary cultured rat hepatocytes and the nuclear fraction, respectively. Lactacystin (5 μM) and epoxomicin (0.5 μM), 26S proteasome inhibitors, significantly suppressed the Dex-dependent maximum increase of TAT and TO mRNA levels in the cells and the nuclear fraction. Electrophoretic mobility shift assay demonstrated that lactacystin did not affect binding of glucocorticoid receptor to glucocorticoid responsive element. Furthermore, lactacystin did not affect the activation of GRE luciferase reporter by Dex transfected to the cells. The results demonstrate that 26S proteasome is positively involved in the Dex-dependent increase of TAT and TO mRNA levels in the cells and suggest that the mechanism of action of 26S proteasome may be degradation of some RNase(s), which breaks down TAT and TO mRNAs.

番木瓜酪氨酸氨基转移酶基因CpTAT的分离与分析

番木瓜酪氨酸氨基转移酶(Tyrosine aminotransferase,TAT)是维生素E合成途径中的一个关键酶。在已有番木瓜果实成熟差异基因片段序列的基础上,采用RACE技术克隆了TAT基因的全长c DNA序列,命名为Cp TAT。该基因全长包含5′非编码区98 bp,3′非编码区232 bp,开放性阅读框1 266 bp,编码421个氨基酸。序列分析表明,该基因为谷草转氨酶超家族成员,催化活性位点为第253位的赖氨酸,编码蛋白与毛果杨、野草莓、桃、拟南芥和水稻的TAT蛋白同源性分别为83.69%、81.09%、80.76%、78.87%、54.67%。荧光定量分析表明,Cp TAT基因在根中的表达量最高,在茎和果实中的表达较低,在叶中的表达量最低。与对照相比,外源乙烯处理能诱导番木瓜果实中Cp TAT基因表达量升高,而1-MCP处理则抑制了该基因表达,在果实衰老期表达量升高,该基因可能在番木瓜果实成熟衰老中起作用。

烟草酪氨酸转氨酶基因家族分析

为揭示烟草酪氨酸转氨酶(Tyrosine aminotransferase,TAT)基因家族的遗传特征和功能,利用生物信息学对不同烟草品种中该基因家族进行鉴定和分析。结果表明,普通烟草中存在4个TATs基因,林烟草和绒毛状烟草中各有3个TATs基因。亚家族Ⅰ中TATs基因的编码序列在1 263~1 338 bp,编码404~445个氨基酸,蛋白质分子量为42.81~49.64 k D,均为不含跨膜结构域的稳定蛋白。亚家族Ⅱ中TATs蛋白均为不稳定蛋白。除NtabTAT4外,其余TATs基因均含有7个外显子和6个内含子。烟草TATs蛋白可分为两类:Group Ⅰ和Group Ⅱ。qPCR分析表明,NtabTATs在普通烟草K326的根、茎、叶和花中都有表达。

紫苏酪氨酸氨基转移酶基因片段的克隆及表达分析

为获得紫苏迷迭香酸合成途径中的酪氨酸氨基转移酶基因,本研究采用同源克隆的方法,根据已报道的其他物种的TAT基因序列设计并合成简并引物,成功克隆得到了紫苏TAT基因片段(GenBank登录号:JN032113.1),该片段长为579bp,共编码193个氨基酸残基,并命名为PfTAT。氨基酸序列比对分析发现其与彩叶草、丹参、拟南芥和罂粟的一致性分别为97%、94%、69%和58%,系统进化树分析表明PfTAT与唇形科植物的亲缘关系最近。采用半定量RT-PCR法分析PfTAT在紫苏的根、茎、叶中均有表达,且叶中的表达量较高,内源性植物激素信号分子对PfTAT表达量影响的实验表明,脱落酸、水杨酸处理均能够不同程度得上调PfTAT转录水平的表达。