Effects of cognitive training on working memory of female methamphetamine-dependent youths / 中华行为医学与脑科学杂志

Objective:To explore the characteristics of working memory ability of female methamphetamine dependent youths, and to analyze the effect of spatial and temporal training task (CODING) on working memory ability of female methamphetamine dependent youths.Methods:Sixty methamphetamine dependent youths from a women's drug rehabilitation center in Sichuan province were selected as the methamphetamine dependent group, while 60 female healthy volunteers from vocational high schools and universities participated as the healthy control group. Visual memory test (VISGED) was used to evaluat the working memory ability of the subjects, and SPSS 21.0 software was used for the nonparametric test of the memory difference between the two groups.The methamphetamine dependent group was divided into treatment group and waiting group based on the baseline matching principle of working memory. The treatment group was trained under the spatio-temporal CODING paradigm for working memory, and the training effect was assessed using the independent sample t-test. Results:(1)The individual parameters(0.06(-1.62, 1.67) vs 1.93 (-0.28, 2.71)) and working program(15.00(14.00 vs 16.00), 17.00(15.00, 20.00)) of working memory in methamphetamine dependent group and normal control group were statistically significant ( Z=-3.83, P<0.05, Z=-3.50; P<0.05). (2) Through memory training, the personal parameters before and after training in the treatment group ((0.08±1.79) vs 1.68 (0.68, 2.25)) had a statistically significant difference ( Z=-3.63, P<0.05). The personal parameters before and after the test in the waiting group ((-0.01±1.86) vs 0.72(-1.54, 1.88)) had no statistically significant difference ( Z=-1.35, P>0.05). Conclusion:Methamphetamine dependence impaire the working memory of female youths regardless of age or education level. Memory training can restore and improve memory ability to a certain extent, help to restore learning ability and living ability, and lay a foundation for a better return to society.

The clinical effect of posterior fossa decompression combined with dural reconstruction in the treatment of Chiari malformation-Ⅰ complicated with syringomyelia / 中国医师进修杂志

Objective:To investigate the clinical effect of posterior fossa decompression combined with dural reconstruction in the treatment of Chiari malformation-Ⅰ(CM-Ⅰ) complicated with syringomyelia (SM).Methods:The clinical data of 50 patients with CM-Ⅰ complicated with SM who were treated in Yan′an University Xianyang Hospital from June 2019 to January 2021 were analyzed. They were divided into the study group (27 cases) and the control group (23 cases) according to the surgical methods. The former received posterior fossa decompression combined with dural reconstruction, while the latter received posterior fossa decompression alone. The clinical symptom improvement, neurological function, cerebrospinal fluid dynamics and syringomyelia changes were compared between the two groups before and after the surgery, and postoperative complications were compared.Results:The overall clinical symptom improvement rate between the two groups had no significant difference ( P> 0.05). After the surgery, the scores of pain, sensory disturbance, dyskinesia and ataxia in the study group were higher than those in the control group: (4.56 ± 0.35) points vs. (4.28 ± 0.43) points, (3.61 ± 0.82) points vs. (3.15 ± 0.73) points, (3.81 ± 0.44) points vs. (3.59 ± 0.50) points, (4.43 ± 0.41) points vs. (4.09 ± 0.53) points, there were statistical significant ( P<0.05). After the surgery, the cerebrospinal fluid stroke volume (SV) and mean flow (MF) in the study group were higher than those in the control group: (0.05 ± 0.02) ml vs. (0.04 ± 0.01) ml, (0.05 ± 0.01) ml/s vs. (0.04 ± 0.01) ml/s; the maximum peak flow velocity (V max) of the head and tail in the study group were lower than those in the control group: (3.14 ± 1.05) mm/s vs. (3.87 ± 1.13) mm/s, (5.56 ± 1.38) mm/s vs. (6.43 ± 1.22) mm/s, there were statistical significant ( P<0.05). There were no significant differences in the rate of reduction or disappearance of syringomyelia, the rate of no change and the rate of increase of syringomyelia after the surgery between the two groups ( P>0.05). There was no significant difference in the incidence of postoperative complications between the two groups ( P>0.05). Conclusions:Posterior fossa decompression combined with dural reconstruction in CM-Ⅰ complicated with SM can better improve cerebrospinal fluid dynamics, and promote the reduction of syringomyelia without increasing postoperative complications.

Effect of cognitive training intervention on attention deficit of methamphetamine-dependent female youth / 中华行为医学与脑科学杂志

Objective:To explore the effect of methamphetamine (MA) dependence on the attention of female youth, and to analyze the intervention effect of computer-based cognitive training on the attention of MA-dependent female youth.Method:From May to August 2021, a total of 64 MA-dependent female youths in abstinence period from a women's compulsory isolated drug rehabilitation center in Sichuan Province were selected as the MA group, and 53 ordinary female youths matched with their age and education level were selected as the normal control group.According to the matching principle of age, education level, and attention pre-test scores, the subjects in MA-dependent group were divided into MA-dependent intervention group( n=30)and MA-dependent waiting group( n=34). CogniPlus cognitive training system was used to train the attention of subjects in MA-dependent intervention group( n=30), while the subjects in MA-dependent waiting group( n=34) and normal control group did not receive training.The Vienna Test System was used to collect the attention scores of all subjects.SPSS 20.0 analysis software was used for data processing.Statistical analysis was performed using independent samples t test and paired samples t test. Results:The MA-dependent group had significantly longer endogenous alertness, temporary alertness, and concentration response time ((275.61±47.79)ms, (268.63±51.41)ms, (444.08±134.40)ms) compared with the normal control group ((247.02±34.09)ms, (237.60±46.04)ms, (355.15±44.37)ms) ( t=3.767, 3.405, 4.976; all P<0.05). After attention training, the post-test reaction time of endogenous alertness, temporary alertness, and concentration ((264.10±38.98)ms, (251.67±38.06)ms, (352.03±65.70)ms) in the MA-dependent intervention group were significantly shorter than those in the MA-dependent waiting group ((323.18±83.28)ms, (302.74±82.75)ms, (402.76±74.34)ms) ( t=-3.702, -3.232, -2.876; all P<0.05). The post-test reaction time of temporary alertness and concentration in the MA-dependent intervention group ((251.67±38.06)ms, (352.03±65.70)ms)were shorter than those of the pre-test ((265.70±37.84)ms, (428.67±120.11)ms) ( t=2.179, 3.588; both P<0.05). The MA-dependent waiting group had a longer post-test reaction time of endogenous alertness and temporary alertness(323.18±83.28)ms, (302.74±82.75)ms) compared with the pre-test ((285.35±51.43)ms, (271.21±61.42)ms) ( t=-2.752, -2.664; both P<0.05) and the post-test reaction time of concentration ((402.76±74.34)ms) was shorter than that of the pre-test ((457.68±146.29)ms)( t=2.431, P<0.05). The MA-dependent intervention group had longer endogenous alertness, temporary alertness, and pre-test reaction time of concentration ((264.57±41.41)ms, (265.70±37.84)ms, (428.67±120.11)ms)compared with the normal control group( t=2.083, 2.841, 3.230; all P<0.05). The post-test of endogenous alertness ((264.10±38.98)ms) was longer than that of the normal control group ( t=2.082, P<0.05). Conclusion:The cognitive training based on the CogniPlus system has a certain effect on the attention maintenance or recovery of MA-dependent female youth.It can be used as an intervention measure for cognitive impairment of drug addicts and help them healthy return to society.

Drift correction in localization microscopy using entropy minimization.

Localization microscopy offers resolutions down to a single nanometer but currently requires additional dedicated hardware or fiducial markers to reduce resolution loss from the drift of the sample. Drift estimation without fiducial markers is typically implemented using redundant cross correlation (RCC). We show that RCC has sub-optimal precision and bias, which leaves room for improvement. Here, we minimize a bound on the entropy of the obtained localizations to efficiently compute a precise drift estimate. Within practical compute-time constraints, simulations show a 5x improvement in drift estimation precision over the widely used RCC algorithm. The algorithm operates directly on fluorophore localizations and is tested on simulated and experimental datasets in 2D and 3D. An open source implementation is provided, implemented in Python and C++, and can utilize a GPU if available.

mRNA structural dynamics shape Argonaute-target interactions.

Small interfering RNAs (siRNAs) promote RNA degradation in a variety of processes and have important clinical applications. siRNAs direct cleavage of target RNAs by guiding Argonaute2 (AGO2) to its target site. Target site accessibility is critical for AGO2-target interactions, but how target site accessibility is controlled in vivo is poorly understood. Here, we use live-cell single-molecule imaging in human cells to determine rate constants of the AGO2 cleavage cycle in vivo. We find that the rate-limiting step in mRNA cleavage frequently involves unmasking of target sites by translating ribosomes. Target site masking is caused by heterogeneous intramolecular RNA-RNA interactions, which can conceal target sites for many minutes in the absence of translation. Our results uncover how dynamic changes in mRNA structure shape AGO2-target recognition, provide estimates of mRNA folding and unfolding rates in vivo, and provide experimental evidence for the role of mRNA structural dynamics in control of mRNA-protein interactions.

High-Speed Super-Resolution Imaging Using Protein-Assisted DNA-PAINT.

Super-resolution imaging allows for the visualization of cellular structures on a nanoscale level. DNA-PAINT (DNA point accumulation in nanoscale topology) is a super-resolution method that depends on the binding and unbinding of DNA imager strands. The current DNA-PAINT technique suffers from slow acquisition due to the low binding rate of the imager strands. Here we report on a method where imager strands are loaded into a protein, Argonaute (Ago), which allows for faster binding. Ago preorders the DNA imager strand into a helical conformation, allowing for 10 times faster target binding. Using a 2D DNA origami structure, we demonstrate that Ago-assisted DNA-PAINT (Ago-PAINT) can speed up the current DNA-PAINT technique by an order of magnitude, while maintaining the high spatial resolution. We envision this tool to be useful for super-resolution imaging and other techniques that rely on nucleic acid interactions.

Argonaute bypasses cellular obstacles without hindrance during target search.

Argonaute (Ago) proteins are key players in both gene regulation (eukaryotes) and host defense (prokaryotes). Acting on single-stranded nucleic-acid substrates, Ago relies on base pairing between a small nucleic-acid guide and its complementary target sequences for specificity. To efficiently scan nucleic-acid chains for targets, Ago diffuses laterally along the substrate and must bypass secondary structures as well as protein barriers. Using single-molecule FRET in conjunction with kinetic modelling, we reveal that target scanning is mediated through loose protein-nucleic acid interactions, allowing Ago to slide short distances over secondary structures, as well as to bypass protein barriers via intersegmental transfer. Our combined single-molecule experiment and kinetic modelling approach may serve as a platform to dissect search processes and study the effect of sequence on search kinetics for other nucleic acid-guided proteins.

DNA-guided DNA cleavage at moderate temperatures by Clostridium butyricum Argonaute.

Prokaryotic Argonaute proteins (pAgos) constitute a diverse group of endonucleases of which some mediate host defense by utilizing small interfering DNA guides (siDNA) to cleave complementary invading DNA. This activity can be repurposed for programmable DNA cleavage. However, currently characterized DNA-cleaving pAgos require elevated temperatures (≥65°C) for their activity, making them less suitable for applications that require moderate temperatures, such as genome editing. Here, we report the functional and structural characterization of the siDNA-guided DNA-targeting pAgo from the mesophilic bacterium Clostridium butyricum (CbAgo). CbAgo displays a preference for siDNAs that have a deoxyadenosine at the 5'-end and thymidines at nucleotides 2-4. Furthermore, CbAgo mediates DNA-guided DNA cleavage of AT-rich double stranded DNA at moderate temperatures (37°C). This study demonstrates that certain pAgos are capable of programmable DNA cleavage at moderate temperatures and thereby expands the scope of the potential pAgo-based applications.

Facilitated diffusion of Argonaute-mediated target search.

Argonaute (Ago) proteins are of key importance in many cellular processes. In eukaryotes, Ago can induce translational repression followed by deadenylation and degradation of mRNA molecules through base pairing of microRNAs (miRNAs) with a complementary target on a mRNA sequence. In bacteria, Ago eliminates foreign DNA through base pairing of siDNA (small interfering DNA) with a target on a DNA sequence. Effective targeting activities of Ago require fast recognition of the cognate target sequence among numerous off-target sites. Other target search proteins such as transcription factors (TFs) are known to rely on facilitated diffusion for this goal, but it is undetermined to what extent these small nucleic acid-guided proteins utilize this mechanism. Here, we review recent single-molecule studies on Ago target search. We discuss the consequences of the recent findings on the search mechanism. Furthermore, we discuss the open standing research questions that need to be addressed for a complete picture of facilitated target search by small nucleic acids.

Characteristics and influencing factors of emotional face recognition on Methamphetamine depend-ence adolescents / 中华行为医学与脑科学杂志

Objective To explore the characteristics and influencing factors of methamphetamine dependence adolescents’ emotional faces recognition ability. Methods Eighty-six adolescents methamphet-amine addicts and 85 healthy volunteers were tested by a self-designed questionnaire and an experiment of e-motional faces identification. Results (1)There was no significant correlation between the emotional faces recognition ability of the experimental group with their age,education level and drug time (P>0. 05). (2)The correct number of positive(32. 58 ±8. 56),neutral (32. 76±8. 06),and negative(56. 28±20. 04) emotional faces recognition in the experimental group was significantly lower than that in the control group of positive (35. 64±3. 91),neutral(35. 47±4. 00),and negative(71. 02±8. 62) emotional faces recognition(t=-3. 00,-2. 79,-6. 24,P<0. 05). Meanwhile,the response time of positive(( 401. 32± 175. 13) ms) and negative ((502. 08±194. 42)ms) emotional faces recognition in the experimental group was significantly lower than that in the control group of positive((300. 83±139. 48)ms) and negative((379. 91±197. 30)ms) emotional faces (t=4. 15,4. 08,P<0. 05). Nevertheless,there was no significant difference between the experimental group((400. 90±174. 21) ms) and the control group ((356. 67±156. 70) ms) in the recognition response time of neutral emotional faces (t=1. 75,P>0. 05). Conclusion Methamphetamine dependence impairs the ability of adolescents’ emotional faces recognition,and their positive and negative emotional recognition abili-ty has processing defects. The ability of methamphetamine dependence adolescents to emotional faces recogni- tion is not affected by their age,education level and time of drug uses.

Study on Protective Effect of Huayu Qutan Decoction on Vascular Dementia / 中国实验方剂学杂志

Objective: To study the protective effect of Huayu Qutan decoction on vascular dementia (VD) gerbils and to explore whether its mechanism is related to Calcium ion-calmodulin-dependent protein kinase Ⅱ (CaMKⅡ)/cyclic adenosine effect element binding protein (CREB)/brain-derived neurotrophic factor (BDNF) signaling pathway. Method: Forty healthy gerbils were randomly divided into sham operation group, model group, low, medium and high dose groups (5.35, 10.7, 21.4 g·kg-1) of removing blood stasis and expelling phlegm. Eight gerbils in each group were divided into model group and removing blood stasis and expelling phlegm group. Gerbils were given corresponding drugs twice a day after operation. Water maze experiment was conducted 21 days later to investigate the spatial learning and memory ability of gerbils. The expression of p-CaMKⅡ/CaMKⅡ, p-CREB/CREB and BDNF in the hippocampus of gerbils were detected by Western blot and immunohistochemistry. Result: Compared with sham operation group, the incubation period and the number of platform trips of gerbil in the model group were significantly reduced, p-CaMKⅡ/CaMKⅡ, p-CREB/CREB, and BDNF protein expression were significantly reduced (PPPConclusion: Huayu Qutan decoction improves the learning and memory abilities of gerbils with vascular dementia, and its mechanism may be related to the activation of CaMKⅡ/CREB/BDNF signaling pathway.

RecA filament maintains structural integrity using ATP-driven internal dynamics.

At the core of homologous DNA repair, RecA catalyzes the strand exchange reaction. This process is initiated by a RecA loading protein, which nucleates clusters of RecA proteins on single-stranded DNA. Each cluster grows to cover the single-stranded DNA but may leave 1- to 2-nucleotide (nt) gaps between the clusters due to three different structural phases of the nucleoprotein filaments. It remains to be revealed how RecA proteins eliminate the gaps to make a seamless kilobase-long filament. We develop a single-molecule fluorescence assay to observe the novel internal dynamics of the RecA filament. We directly observe the structural phases of individual RecA filaments and find that RecA proteins move their positions along the substrate DNA to change the phase of the filament. This reorganization process, which is a prerequisite step for interjoining of two adjacent clusters, requires adenosine triphosphate hydrolysis and is tightly regulated by the recombination hotspot, Chi. Furthermore, RecA proteins recognize and self-align to a 3-nt-period sequence pattern of TGG. This sequence-dependent phase bias may help the RecA filament to maintain structural integrity within the kilobase-long filament for accurate homology search and strand exchange reaction.

Clinical Application of Detecting Serum PPAR-γ mRNA,MMP-9 mRNA in the Diagnosis of Ruptured Intracranial Aneurysm / 现代检验医学杂志

Objective To investigate the clinical application of detecting serum PPAR-γmRNA,MMP-9mRNA in the diagnosis of ruptured intracranial aneurysm.Methods The expression of serum PPAR-γmRNA,MMP-9mRNA were detected for 87 cases of patients with intracranial aneurysm,including ruptured group and non-ruptured group,respectively,with 57 cases and 30 cases of patients.The control group should be established to compare the changes of the above indicators.Results The expression of serum PPAR-γmRNA in the ruptured group,the non-ruptured and the controls group were 0.23±0.03,0.59±0.11 and 0.87±0.15,which of MMP-9mRNA were 0.93±0.17,0.63±0.13 and 0.25±0.05.Compared with those in the controls group,the expression of serum PPAR-γmRNA in the ruptured group significantly lowered (t=23.79,P＜0.01),which of MMP-9mRNA raised (t=25.63,P＜0.01).There were statistically significant differences.The expression of PPAR γmRNA in the ruptured group were lower than those in the unruptured group,which of MMP-9 mRNA were higher (t=15.32,16.27,P＜0.01).To establishing the receiver-operating characteristic curve (ROC curve) in evaluating the clinical significances of the two markers to use the rupture group and non-ruptured group as the dependent variable,the AUC of the expression of serum PPAR-ymRNA,MMP-9mRNA were 0.858 (95 ％ CI:0.775 ～ 0.940,P =0.000),0.842 (95 ％CI:0.756～0.929,P=0.000).As the dependent variable in the control group and unruptured group,the AUC of the expression of serum PPAR-γmRNA,MMP-9mRNA were 0.827 (95％CI:0.734～0.920,P=0.000);0.818 (95％CI:0.722 ～0.914,P=0.000).Conclusion Detection of serum PPAR-γ mRNA,MMP-9 mRNA can be applied in assessment of occurrence and progression for the intracranial aneurysm,and to provide evidences for the early detection of ruptured intracranial aneurysm.

High Spatiotemporal-Resolution Magnetic Tweezers: Calibration and Applications for DNA Dynamics.

The observation of biological processes at the molecular scale in real time requires high spatial and temporal resolution. Magnetic tweezers are straightforward to implement, free of radiation or photodamage, and provide ample multiplexing capability, but their spatiotemporal resolution has lagged behind that of other single-molecule manipulation techniques, notably optical tweezers and AFM. Here, we present, to our knowledge, a new high-resolution magnetic tweezers apparatus. We systematically characterize the achievable spatiotemporal resolution for both incoherent and coherent light sources, different types and sizes of beads, and different types and lengths of tethered molecules. Using a bright coherent laser source for illumination and tracking at 6 kHz, we resolve 3 Å steps with a 1 s period for surface-melted beads and 5 Å steps with a 0.5 s period for double-stranded-dsDNA-tethered beads, in good agreement with a model of stochastic bead motion in the magnetic tweezers. We demonstrate how this instrument can be used to monitor the opening and closing of a DNA hairpin on millisecond timescales in real time, together with attendant changes in the hairpin dynamics upon the addition of deoxythymidine triphosphate. Our approach opens up the possibility of observing biological events at submillisecond timescales with subnanometer resolution using camera-based detection.

GATA5 loss-of-function mutations associated with congenital bicuspid aortic valve.

Bicuspid aortic valve (BAV) is the most common form of congenital cardiovascular defect in humans worldwide and is responsible for substantial morbidity and mortality. Accumulating evidence has demonstated that genetic risk factors are involved in the pathogenesis of BAV. However, BAV is genetically heterogeneous and the genetic basis underlying BAV in a large number of patients remains unknown. In the present study, the coding regions and splice junction sites of the GATA5 gene, which codes for a zinc-finger transcription factor crucial for the normal development of the aortic valve, was sequenced initially in 110 unrelated patients with BAV. The available relatives of the mutation carriers and 200 unrelated healthy individuals used as controls were subsequently genotyped for GATA5. The functional effect of the mutations was characterized by using a luciferase reporter assay system. As a result, two novel heterozygous GATA5 mutations, p.Y16D and p.T252P, were identified in two families with autosomal dominant inheritance of BAV, respectively. The variations were absent in 400 control chromosomes and the altered amino acids were completely conserved evolutionarily. Functional assays revealed that the two GATA5 mutants were associated with significantly reduced transcriptional activity compared with their wild-type counterpart. To the best of our knowledge, this is the first study on the association of GATA5 loss-of-function mutations with enhanced susceptibility to BAV, providing novel insight into the molecular mechanism involved in human BAV and suggesting a potential role for the early prophylaxis and personalized treatment of this common congenital heart disease.

[The ultrastructural features of sputum deposition and its value in the diagnosis of pulmonary alveolar proteinosis].

OBJECTIVE: To investigate the ultrastructural features of sputum deposition (SD) and its value in the diagnosis of pulmonary alveolar proteinosis (PAP). METHODS: Seven patients with PAP diagnosed by lung biopsy and cytology were enrolled in this study. The patients consisted of 5 men and 2 women, whose median age was 48 years (range 36 to 73). SD and bronchoalveolar lavage fluid (BALF) sediment were made into ultrathin sections and observed under transmission electron microscope (TEM), respectively. Seven cases of control group composed of 4 men and 3 women whose median age was 49 years (range 39 to 68) including 3 cases of bacterial pneumonia, two cases of COPD and 2 cases of exudative pulmonary tuberculosis. Each SD was made into ultrathin section, and compared with the experimental group. RESULTS: In PAP group, Periodic acid-schiff (PAS) staining was performed on 7 sputum smears and none of them was tested positive for any components with diagnostic interest. Four cases from the 7 paraffin-embed sections of BALF sediment by microscopic examination suggested PAP. Under TEM, BALF sediment showed that many lamellar bodies existed in and outside alveolar epithelial cells, and 5 specimens were consistent with PAP diagnosis. Compared with BALF sediment, SD had apparent degeneration with more myelin phagosomes in the cytoplasm of macrophages, more lamellar bodies in alveolar epithelial cells, and lots of lamellar bodies in the shape of concentric circle in the extracellular spaces. Four from the 7 SD samples were consistent with the diagnosis of PAP. No significant difference was found between SD and BALF in the diagnosis of PAP by electronic examination (P > 0.05). In the 7 cases of control group no drifting osmiophilic lamellar bodies in extracellular space were detected. CONCLUSIONS: The osmiophilic lamellar bodies with diagnostic value were found in SD and BALF of patients with PAP. TEM of SD in combination with clinical manifestations and radiologic findings can make a definitive diagnosis of PAP, especially for those patients who have contraindications to lung biopsy and lung lavage.

Synthesis, structure, molecular docking, and structure-activity relationship analysis of enamines: 3-aryl-4-alkylaminofuran-2(5H)-ones as potential antibacterials.

Thirty-one 3-aryl-4-alkylaminofuran-2(5H)-ones were designed, prepared and tested for their antibacterial activity. Some of them showed significant antibacterial activity against Gram-positive organisms, especially against Staphylococcus aureus ATCC 25923, but all were inactive against Gram-negative organisms. Out of these compounds, 3-(4-bromophenyl)-4-(2-(4-nitrophenyl)hydrazinyl)furan-2(5H)-one (4a11) showed the most potent antibacterial activity against S. aureus ATCC 25923 with MIC(50) of 0.42 µg/mL. The enzyme assay revealed that the possible antibacterial mechanism of the synthetic compounds might be due to their inhibitory activity against tyrosyl-tRNA synthetase. Molecular dockings of 4a11 into S. aureus tyrosyl-tRNA synthetase active site were also performed. This inhibitor snugly fitting the active site might well explain its excellent inhibitory activity. Meanwhile, this modeling disclosed that a more suitable optimization strategy might be to modify the benzene ring at 3-position of furanone with hydrophilic groups.

Binding orientations, QSAR, and molecular design of thiophene derivative inhibitors.

A theoretical study on binding orientations and quantitative structure-activity relationship of thiophene derivatives as inhibitors towards tubulin has been carried out by using the docking analysis and the comparative molecular field analysis. The appropriate binding orientations and conformations of these compounds interacting with tubulin were revealed by docking study; and a 3D-quantitative structure-activity relationship model showing significant statistical quality and satisfying predictive ability was established, in which the correlation coefficient (R(2)) and cross-validation coefficient (q(2)) are 0.949 and 0.743, respectively. The same model was further applied to predict the pIC(50) values for nine congeneric compounds as external test set, and the predictive correlation coefficient R(pred)(2) reaches 0.929, thus the predictive ability of this 3D-quantitative structure-activity relationship model can be further confirmed. Some key structural factors of the compounds responsible for cytotoxicity were discussed in detail. Based on these structural factors, three new compounds with higher activity have been designed, and their cytotoxicities were also predicted by the established 3D-quantitative structure-activity relationship model from comparative molecular field analysis as well as the docking analysis. We hope these theoretical results can be confirmed by experimental work.

Expression differentiation of CYC-like floral symmetry genes correlated with their protein sequence divergence in Chirita heterotricha (Gesneriaceae).

CYCLOIDIEA (CYC) and its homologues have been studied intensively in the model organism Antirrhinum majus and related species regarding their function in controlling floral dorsoventral (adaxial-abaxial) asymmetry, including aborting the adaxial and lateral stamens. This raises the question whether the same mechanism underlies the great morphological diversity of zygomorphy in angiosperms, especially in Lamiales sensu lato, a major clade predominantly with zygomorphic flowers. To address this, we selected a representative in Gesneriaceae, the sister to the remainder of Lamiales s.l., to isolate CYC homologues and further investigate their expression patterns using locus-specific semiquantitative reverse transcriptase polymerase chain reaction. Our results showed that four CYC homologues in Chirita heterotricha differentiated spatially and temporally in expression, in which ChCYC1D was only expressed in the adaxial regions, and transcripts of ChCYC1C were distributed in both the adaxial and lateral regions, while ChCYC2A and ChCYC2B transcripts were only detected in the young inflorescences. ChCYC1C expression in the lateral regions correlated with abortion of the lateral stamens in C. heterotricha hinted at its gain of function, i.e., expanding from the adaxial to the lateral regions in expression. Correlatively, the protein sequences of ChCYC genes exhibited remarkable divergences, in which some lineage-specific amino acids between GCYC1 and GCYC2 in conserved functional domains and two sublineage-specific motifs between GCYC1C and GCYC1D in GCYC1 genes had further been identified. Our results indicated that ChCYC genes had probably undergone an expressional differentiation and specialization in establishing the floral dorsoventral asymmetry in C. heterotricha responding to different selective pressure after gene duplication.