Identification, characterization, and expression analysis of WRKY transcription factors in Cardamine violifolia reveal the key genes involved in regulating selenium accumulation.

BACKGROUND: Cardamine violifolia is a significant Brassicaceae plant known for its high selenium (Se) accumulation capacity, serving as an essential source of Se for both humans and animals. WRKY transcription factors play crucial roles in plant responses to various biotic and abiotic stresses, including cadmium stress, iron deficiency, and Se tolerance. However, the molecular mechanism of CvWRKY in Se accumulation is not completely clear. RESULTS: In this study, 120 WRKYs with conserved domains were identified from C. violifolia and classified into three groups based on phylogenetic relationships, with Group II further subdivided into five subgroups. Gene structure analysis revealed WRKY variations and mutations within the CvWRKYs. Segmental duplication events were identified as the primary driving force behind the expansion of the CvWRKY family, with numerous stress-responsive cis-acting elements found in the promoters of CvWRKYs. Transcriptome analysis of plants treated with exogenous Se and determination of Se levels revealed a strong positive correlation between the expression levels of CvWRKY034 and the Se content. Moreover, CvWRKY021 and CvWRKY099 exhibited high homology with AtWRKY47, a gene involved in regulating Se accumulation in Arabidopsis thaliana. The WRKY domains of CvWRKY021 and AtWRKY47 were highly conserved, and transcriptome data analysis revealed that CvWRKY021 responded to Na2SeO4 induction, showing a positive correlation with the concentration of Na2SeO4 treatment. Under the induction of Na2SeO3, CvWRKY021 and CvWRKY034 were significantly upregulated in the roots but downregulated in the shoots, and the Se content in the roots increased significantly and was mainly concentrated in the roots. CvWRKY021 and CvWRKY034 may be involved in the accumulation of Se in roots. CONCLUSIONS: The results of this study elucidate the evolution of CvWRKYs in the C. violifolia genome and provide valuable resources for further understanding the functional characteristics of WRKYs related to Se hyperaccumulation in C. violifolia.

[Analysis of SCA3/MJD3 gene mutation and genetic polymorphism in a guangxi family with spinocerebellar ataxia 3].

Autosomal dominant cerebellar ataxias (ADCAs) comprise a group of genetically heterogeneous neurodegenerative disorders among which spinocerebellar ataxia type 3 (SCA3) represents the most common form of SCAs worldwide. The fragments of SCA3/MJD gene,which is the member of family GXPL1,were amplified by polymerase chain reaction (PCR). The PCR products of SCA3/MJD gene were detected with capillary electrophoresis (CE) and sequencing to evaluate the size of CAG repeats, feature in the transmission and the mutation in the family with SCA3 in Guangxi province. The results showed that the exon 10 of the SCA3/MJD gene contains 64-71 CAG repeats in all of the affected individuals and three asymptomatic carriers of the family. The number of the CAG repeats during transmission in the normal individuals carrying CGG allele remains consistent, suggesting that CGG allele could have no effect on intergenerational stability of CAG repeats in normal individuals. In addition, two novel point mutations were identified: IVS9-113 T > C in the intronic region and a missense mutation 220 G > A (Glu > Gly) in the encoding region. These two novel point mutations have not been reported and the effect of the mutations on the phenotype of SCA3 is not clear.

[Haplotype analysis for mucocutaneous venous malformations in a Chinese Han ethnic family].

A Chinese Han ethnic family with mucocutaneous venous malformations (VMCM) was investigated. This family has autosomal dominantly inherited VMCM in five generations, and the offspring has a 50% risk of this inherited disorder. Affected individuals have small, spongy, and multiple vascular lesions, which often locate in the skin, oral mucosa, and upper and lower extremities. None of the family members had gastrointestinal bleeding, central nervous system involvement and cardiac defects. Pathological analysis showed that the veins have irregular vascular space and walls with variable thickness. All phenotypes of the patients displayed the basic characters of VMCM. To analyze the genetic locus and haplotype, genomic DNA of 26 family members was obtained from peripheral leukocytes, and the linkage analysis and haplotypes analysis were performed using microsatellites markers. The results of two-point linkage analysis and haplotype analysis showed that the disease-causing gene located within a 7 cM region between D9S1121 and D9S161 on the short arm of chromosome 9. The study firstly reported the Chinese family with VMCM, which disease-causing gene is located in 9p, consistent with western VMCM families reported. Four flanking markers, D9S1121, D9S169, D9S16 and D9S248, were used to define the linkage haplotypes in the family, which can provide useful informaion for researchers to study VMCM in different racial background.

[Retinal cell apoptosis, microvascular changes and expression of connective tissue growth factor in experimental diabetic rats].

OBJECTIVE: To study the cell apoptosis and the expression of connective tissue growth factor (CTGF) in the retina of diabetic rats and to explore their contributions to the changes of microcirculation. METHODS: It was a experiment study. Fifty-five adult male Wistar rats were divided into two groups, normal control group (CON, 10 rats) and diabetes mellitus group (DM, 45 rats). The 30 surviving rats in the DM group were further divided into 3 groups based on the time of observation, 2 month (DM2), 4 month (DM4) and 6 month (DM6) groups, with 10 rats in each group. Cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL). Expression of CTGF was determined by immunohistochemical study. Retinal vessels were observed by retinal digest stretched periodic acid Schiff (PAS) staining. RESULTS: Immunohistochemical staining and TUNEL staining revealed negative results in normal control group. Retinal cell apoptosis index increased gradually from DM2 to DM6, the differences between any two groups were statistically significant (t(2-4, 2-6, 4-6) = 21.432, 50.843, 29.410; P < 0.05). Expression of CTGF in the retina increased from DM2-DM6, the differences between any two groups were statistically significant (t(2-4, 2-6, 4-6) = 15.345, 26.316, 10.971; P < 0.05). PAS staining of retinal blood vessels obtained negative results in the CON and DM2 groups. Part of retinal capillaries were slightly stiff and narrow in DM4 group. Retinal capillaries in DM6 group were trunk stiff and were narrowed obviously. The number of pericytes was reduced in DM4, and progressed following the course of diabetes. The number of pericytes in the DM2 group did not different from that in the CON group (t = 0.875, P = 0.387). The number of pericytes in the DM4 and DM6 group were significantly decreased as compared to the CON group (t = 3.367, 6.667; P < 0.05). Retinal cellular apoptosis index had a significant positive correlation to the expression of CTGF (r = 0.958, P < 0.05). Number of pericytes was significantly correlated (negative correlation) with retinal cellular apoptosis index and the expression of CTGF (r = -0.540, -0.595; P < 0.05). CONCLUSIONS: The appearances of cellular apoptosis and fibrosing factor CTGF in the retina of diabetic rats occurred earlier than the changes of microcirculation and the number of capillary pericytes.

[Gene diagnosis and CAG repeat analysis of spinocerebellar ataxia cases of Guangxi region].

To characterize the distributions and subtypes of the spinocerebellar ataxias (SCA) in Guangxi region, the SCAl, SCA2, SCA3/MJD, SCA6, SCA7 and SCA12 (CAG)n mutations were analyzed by polymerase chain reaction (PCR) and capillary electrophoresis (CE). The SCA3/MJD mutation was detected in a total of 21 SCA patients and 19 presymptomatic individuals from 6 SCA families and their CAG repeat numbers were 59-70 and 60-73, respectively. No (CAG)n mutations of SCA1, SCA2, SCA6, SCA7 and SCA12 were detected. This study showed that SCA in Guangxi region is mostly SCA3/MJD subtype and the CAG repeats are smaller than those reported in other regions previously.

[Observation on polytene chromosomes of the ovarian nurse cell of anopheles minimus from Yuanjiang].

Anopheles minimus collected from Yuanjiang, Yunnan Province, were bred with standard methods in lab. The ovarian nurse cells of A.minimus were separated and stained, and the whole polytene chromosomes were photographed under light microscope and compared with A.minimus from Guangxi. 365 samples of ovarian nurse cells were observed. The chromosomes included one telocentric sex-chromosome X, two submetacentric autosomes II (autosome II right arm, 2R and autosome II left arm, 2L) and two metacentric autosomes III (autosome III right arm, 3R, and autosome III left arm, 3L). The X is the shortest chromosome and the 2R is the longest one. In comparison with the pattern of polytene chromosomes of A. minimus from Guangxi, difference at 12 positions has been found at the parts of arms in banding sequences.

Ginsenoside R(e) increases fertile and asthenozoospermic infertile human sperm motility by induction of nitric oxide synthase.

We investigated the effects of Ginsenoside R(e) on human sperm motility in fertile and asthenozoospermic infertile individuals in vitro and the mechanism by which the Ginsenosides play their roles. The semen samples were obtained from 10 fertile volunteers and 10 asthenozoospermic infertile patients. Spermatozoa were separated by Percoll and incubated with 0, 1, 10 or 100 microM of Ginsenoside R(e). Total sperm motility and progressive motility were measured by computer-aided sperm analyzer (CASA). Nitric oxide synthase (NOS) activity was determined by the 3H-arginine to 3H-citrulline conversion assay, and the NOS protein was examined by the Western blot analysis. The production of sperm nitric oxide (NO) was detected using the Griess reaction. The results showed that Ginsenoside R(e) significantly enhanced both fertile and infertile sperm motility, NOS activity and NO production in a concentration-dependent manner. Sodium nitroprusside (SNP, 100 nM), a NO donor, mimicked the effects of Ginsenoside R(e). And pretreatment with a NOS inhibitor N(omega)-Nitro-L-arginine methyl ester (L-NAME, 100 microM) or a NO scavenger N-Acetyl-L-cysteine (LNAC, 1 mM) completely blocked the effects of Ginsenoside R(e). Data suggested that Ginsenoside R(e) is beneficial to sperm motility, and that induction of NOS to increase NO production may be involved in this benefit.

[Expression of target gene in eukaryotic cells driven by prokaryotic T7 promoter and its RNA polymerase].

To enhance the efficiency of the expression of target gene in eukaryotic cells, one of the strongest prokaryotic expression systems, the T7 RNA polymerase and T7 promoter, was introduced into eukaryotic cells. A duel-plasmid gene expression system of T7 bacteriophage components was developed; one containing the T7 phage RNA polymerase gene under the control of eukaryotic promoter CMV (pCMV-T7pol) and the other (pT7IRES) containing the T7 promoter and T7 terminator as well as EMCV IRES. To test the feasibility of this plasmid system for eukaryotic expression, hepatitis B virus envelop HBV preS2/S was used to construct pT7IRES-HBs. The target genes were expressed efficiently by the eukaryonized prokaryotic expression system in a variety of the cells indicating C2C12, SP2/0, NIH3T3 and BALB/c 3T3, suggesting the potential applications of the expression system in gene therapy and gene immunization.

Molecular phylogeny of the genus Paramesotriton (Caudata: Salamandridae).

To elucidate the phylogeny of the genus Paramesotriton (Caudata: Salamandridae), we investigated three mitochondrial DNA gene fragments (1207 bp in total) of cytochrome b, ND2, and ND4 for its six recognized species. The phylogenetic relationships within Paramesotriton were reconstructed by maximum parsimony (MP) and maximum likelihood (ML) methods. Phylogenetic trees (MP and ML trees) that were constructed from the combined data set of the three gene fragments indicated that all six species of Paramesotriton formed a monophyletic group, with P. caudopunctatus as basal to the other five species. This result suggests that P. fuzhongensis is a valid species in Paramesotriton.

Genetic relationship of Chinese ethnic populations revealed by mtDNA sequence diversity.

The origin and demographic history of the ethnic populations of China have not been clearly resolved. In this study, we examined the hypervariable segment I sequences (HVSI) of the mitochondrial DNA control region in 372 individuals from nine Chinese populations and one northern Thai population. A relatively high percentage of individuals was found to share sequences with those from other populations of the same ethnogenesis. In general, the populations of southern or Pai-Yuei tribal origin showed high haplotype diversity and nucleotide diversity compared with the populations of northern or Di-Qiang tribal origin. Mismatch distributions from these populations showed concordant features. All except the northern groups Nu, Lisu, Tibetan, and Mongolian showed typical signatures of ancient population expansions in the mismatch distributions and neutrality tests. Episodes of extreme size reduction in the past are one of the likely explanations for the absence of evidence of expansion in northern populations. Small sample sizes as well as samples from isolated subpopulations contributed to the bumpy mismatch distributions observed. Phylogenetic analysis and haplotype sharing among populations suggest that current mtDNA variation in these ethnic populations could reveal their ethnohistory to some extent, but in general, linguistic and geographic classifications of the populations did not agree well with classification by mtDNA variation.