An Efficient Rice Mutagenesis System Based on Suspension-Cultured Cells

Plant mutants are important bio-resources for crop breeding and gene functional studies. Conventional methods for generating mutant libraries by mutagenesis of seeds with physical or chemical agents are of low efficiency. Here, we developed a highly-efficient ethyl methanesulfonate (EMS) mutagenesis system based on suspension-cultured cells, with rice (Oryza sativa L.) as an example. We show that treatment of suspension-cultured tiny cell clusters with 0.4% EMS for 18-22h followed by differentiation and regeneration produced as high as 29.4% independent mutant lines with visible phenotypic variations, including a number of important agronomic traits such as grain size, panicle size, grain or panicle shape, tiller number and angle, heading date, male sterility, and disease sensitivity. No mosaic mutant was observed in the mutant lines tested. In this mutant library, we obtained a mutant with an abnormally elongated uppermost internode. Sequencing and functional analysis revealed that this is a new allelic mutant of eui (elongated uppermost internode) caused by two point mutations in the first exon of the EUI gene, representing a successful example of this mutagenesis system.     

Tortuosity of the superficial femoral artery and its influence on blood flow patterns and risk of atherosclerosis

Biomechanics and Modeling in Mechanobiology - The superficial femoral artery (SFA) is a typical atherosclerosis-prone site. We aimed to explore whether the tortuosity of the SFA associates with the...     

Sequences downstream of the 5' splice donor site are required for both packaging and dimerization of human immunodeficiency virus type 1 RNA

Two copies of human immunodeficiency virus type 1 RNA are incorporated into each virus particle and are further converted to a stable dimer as the virus particle matures. Several RNA segments that flank the 5' splice donor site at nucleotide (nt) 289 have been shown to act as packaging signals. Among these, RNA stem-loop 1 (SL1) (nt 243 to 277) can trigger RNA dimerization through a "kissing-loop" mechanism and thus is termed the dimerization initiation site. However, it is unknown whether other packaging signals are also needed for dimerization. To pursue this subject, we mutated stem-loop 3 (SL3) (nt 312 to 325), a GA-rich region (nt 325 to 336), and two G-rich repeats (nt 363 to 367 and nt 405 to 409) in proviral DNA and assessed the effects on RNA dimerization by performing native Northern blot analyses. Our results show that the structure but not the specific RNA sequence of SL3 is needed not only for efficient viral RNA packaging but also for dimerization. Mutations of the GA-rich sequence severely diminished viral RNA dimerization as well as packaging; the combination of mutations in both SL3 and the GA-rich region led to further decreases, implying independent roles for each of these two RNA motifs. Compensation studies further demonstrated that the RNA-packaging and dimerization activity of the GA-rich sequence may not depend on a putative interaction between this region and a CU repeat sequence at nt 227 to 233. In contrast, substitutions in the two G-rich sequences did not cause any diminution of viral RNA packaging or dimerization. We conclude that both the SL3 motif and GA-rich RNA sequences, located downstream of the 5' splice donor site, are required for efficient RNA packaging and dimerization.     

Intratumoral expression of IL-17 and its prognostic role in gastric adenocarcinoma patients

In this study, we characterized the intratumoral expression of IL-17 and CD8(+) TILs in gastric adenocarcinoma patients after resection and determined the correlation between the survival probability of gastric adenocarcinoma patients and the expression of IL-17 in tumor. Expression of IL-17 and CD8 was assessed by immunohistochemistry, and the prognostic effects of intratumoral IL-17 expression and CD8(+) TILs were evaluated by Cox regression and Kaplan-Meier analysis. Immunohistochemical detection revealed the presence of IL-17 and CD8(+) cells in gastric adenocarcinoma tissue samples (90.6%, 174 out of 192 patients and 96.9%, 186 out of 192 patients, respectively). We have also found that intratumoral IL-17 expression was significantly correlated with age (p=0.004) and that the number of CD8(+)TILs was significantly correlated with UICC staging (p=0.012) and the depth of tumor invasion (p=0.022). The five-year overall survival probability among patients intratumorally expressing higher levels of IL-17 was significantly better than those expressing lower levels of IL-17 (p=0.036). Multivariate Cox proportional hazard analyses revealed that intratumoral IL-17 expression (HR: 0.521; 95% CI: 0.329-0.823; p=0.005) was an independent factor affecting the five-year overall survival probability. We conclude that low levels of intratumoral IL-17 expression may indicate poor prognosis in gastric adenocarcinoma patients.