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排序方式: 共有1134条查询结果,搜索用时 15 毫秒
1.
Comment on: Rokavec M, et al. Mol Cell 2012; 45:777-89. 相似文献
2.
《Journal of molecular biology》2021,433(17):166665
Transient receptor potential melastatin 4 (TRPM4) is a broadly expressed Ca2+ activated monovalent cation channel that contributes to the pathophysiology of several diseases.For this study, we generated stable CRISPR/Cas9 TRPM4 knockout (K.O.) cells from the human prostate cancer cell line DU145 and analyzed the cells for changes in cancer hallmark functions. Both TRPM4-K.O. clones demonstrated lower proliferation and viability compared to the parental cells. Migration was also impaired in the TRPM4-K.O. cells. Additionally, analysis of 210 prostate cancer patient tissues demonstrates a positive association between TRPM4 protein expression and local/metastatic progression. Moreover, a decreased adhesion rate was detected in the two K.O. clones compared to DU145 cells.Next, we tested three novel TRPM4 inhibitors with whole-cell patch clamp technique for their potential to block TRPM4 currents. CBA, NBA and LBA partially inhibited TRPM4 currents in DU145 cells. However, none of these inhibitors demonstrated any TRPM4-specific effect in the cellular assays.To evaluate if the observed effect of TRPM4 K.O. on migration, viability, and cell cycle is linked to TRPM4 ion conductivity, we transfected TRPM4-K.O. cells with either TRPM4 wild-type or a dominant-negative mutant, non-permeable to Na+. Our data showed a partial rescue of the viability of cells expressing functional TRPM4, while the pore mutant was not able to rescue this phenotype. For cell cycle distribution, TRPM4 ion conductivity was not essential since TRPM4 wild-type and the pore mutant rescued the phenotype.In conclusion, TRPM4 contributes to viability, migration, cell cycle shift, and adhesion; however, blocking TRPM4 ion conductivity is insufficient to prevent its role in cancer hallmark functions in prostate cancer cells. 相似文献
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Gordon Tollin 《Journal of bioenergetics and biomembranes》1995,27(3):303-309
Photoexcitation of flavin analogs generates the lowest triplet state (via intersystem crossing from the first excited singlet state) in the nanosecond time domain and with high quantum efficiency. The triplet, being a strong oxidant, can abstract a hydrogen atom (or an electron) from a reduced donor in a diffusion-controlled reaction. If the donor is a redox protein, the oxidation process can be used to initiate an electron transfer sequence involving either intramolecular or intermolecular reactions. If the donor is an organic compound such as EDTA, the neutral flavin semiquinone will be produced by H atom abstraction; this is a strong reductant and can subsequently transfer a hydrogen atom (or an electron) to an oxidized redox protein, thereby again initiating a sequence of intramolecular or intermolecular processes. If flavin photoexcitation is accomplished using a pulsed laser light source, the initiation of these protein electron transfer reactions can be made to occur in the nanosecond to microsecond time domain, and the sequence of events can be followed by time-resolved spectrophotometry to obtain rate constants and thus mechanistic information. The present paper describes this technology, and selected examples of its use in the investigation of redox protein mechanisms are given. 相似文献
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Elaine C. Kellogg Stephen Kolaczkowski Michael R. Wasielewski David M. Tiede 《Photosynthesis research》1989,22(1):47-59
We have measured the extent of flash-induced electron transfer from the bacteriochlorophyll dimer, P, to the bacteriopheophytin in the M-subunit, HM, in reaction centers of Rhodopseudomonas viridis. This has been done by measuring the transient states produced by excitation of reaction centers trapped in the PHL
–HM state at 90 K. Under these conditions the normal forward electron transfer to the bacteriopheophytin in the L-subunit, HL, is blocked and the yield of transient P+HM
– can be estimated with respect to the lifetime of P*. Under these conditions flash induced absorbance decreases of the bacteriochlorophyll dimer 990 nm band suggest that a transient P+ state is formed with a quantum yield of 0.09±0.06 compared to that formed during normal photochemistry. These transient measurements provide an upper limited on the yield of a transient P+ HM
– state. An estimate of 0.09 as the yield of the P+ HM
– state is consistent with all current observations. This estimate and the lifetime of P* suggest that the electron transfer rate from P* to HM, kM, is about 5 × 109 sec–1 (M = 200ps). These measurements suggest that the a branching ratio kL/kM is on the order of 200. The large value of the branching ratio is remarkable in view of the structural symmetry of the reaction center. This measurement should be useful for electron transfer calculations based upon the reaction center structure. 相似文献
7.
Kees W. Rodenburg Marcel J. A. de Groot Rob A. Schilperoort Paul J. J. Hooykaas 《Plant molecular biology》1989,13(6):711-719
In relation to the question which DNA form (single- or double-stranded) is transferred by Agrobacterium tumefaciens to plant cells, we studied the behaviour of single-stranded DNA, as compared to double-stranded DNA, when it is introduced into plant protoplasts by electroporation. To this end, we cloned a construct with a plant NPTII gene as well as a CAT gene in the M13 vectors tg130 and tg131. We found that both complementary single-stranded molecules gave rise to substantial CAT activity in plant protoplasts, suggesting that single-stranded DNA is converted into double-stranded DNA by the plant cell replication machinery. Unexpectedly, we found that single-stranded DNA leads to a 3–10 fold higher frequency of stable transformation (selection for kanamycin resistance) than double-stranded DNA. These results indicate that the use of single-stranded DNA might be considered in experiments in which optimal transformation frequencies are needed, e.g. with protoplasts form recalcitrant plant species.Abbreviations ss
single-stranded
- ds
double-stranded
- CAT
chloramphenicol acetyl transferase
- NPTII
neomycin phosphotransferase II
- RT
room temperature 相似文献
8.
9.
Bhuvana Gopalakrishnan Burachai Sonthayanon Rownak Rahmatullah Subbaratnam Muthukrishnan 《Plant molecular biology》1991,16(3):463-467
Protoplasts were prepared from barley aleurone layers using Onozuka cellulase digestion and purification through a Percoll gradient. Protoplasts prepared by this procedure had a viability ranging from 60% to 80% during the first two days of culture. They were responsive to gibberellic acid (GA) as measured by the stimulation of -amylase synthesis. The GA stimulation was counteracted by abscisic acid (ABA). In the presence of polyethylene glycol (PEG), the protoplasts took up exogenously added plasmid DNA containing the reporter gene coding for chloramphenicol acetyl transferase (CAT) linked to a 35S promoter from cauliflower mosaic virus (CaMV) or to barley -amylase gene promoters and expressed CAT activity. Therefore, barley aleurone layer protoplasts are suitable for analysis of hormoneresponsive elements in hydrolase genes. 相似文献
10.
Gene structure and expression of a tobacco endochitinase gene in suspension-cultured tobacco cells 总被引:8,自引:0,他引:8
We have isolated and characterized the genomic clone CHN50 corresponding to tobacco basic endochitinase (E.C.3.2.1.14). DNA sequence and blotting analysis reveal that the coding sequence of the gene present on CHN50 is identical to that of the cDNA clone pCHN50 and, moreover, the CHN50 gene has its origin in the progenitor of tobacco, Nicotiana sylvestris. Tobacco basic chitinases are encoded by a small gene family that consists of at least two members, the CHN50 gene and a closely related CHN17 gene which was characterized previously. By northern blot analysis, it is shown that the CHN50 gene is highly expressed in suspension-cultured tobacco cells and the mRNA accumulates at late logarithmic growth phase. To identify cis-DNA elements involved in the expression of the CHN50 gene in suspensioncultured cells, the chimeric gene consisting of 1.1 kb CHN50 5 upstream region fused to the coding sequence of -glucuronidase (GUS) was introduced by electroporation into protoplasts isolated from suspension-cultured tobacco cells. Transient GUS activity was found to be dependent on the growth phase of the cultured cells, from which protoplasts had been prepared. Functional analysis of 5 deletions suggests that the distal region between -788 and -345 contains sequences that potentiate the high-level expression in tobacco protoplasts and the region (-68 to -47) proximal to the TATA box functions as a putative silencer. 相似文献