A comprehensive evaluation of a typical plant telomeric G-quadruplex (G4) DNA reveals the dynamics of G4 formation, rearrangement, and unfolding.

Telomeres are specific nucleoprotein structures that are located at the ends of linear eukaryotic chromosomes and play crucial roles in genomic stability. Telomere DNA consists of simple repeats of a short G-rich sequence: TTAGGG in mammals and TTTAGGG in most plants. In recent years, the mammalian telomeric G-rich repeats have been shown to form G-quadruplex (G4) structures, which are crucial for modulating telomere functions. Surprisingly, even though plant telomeres are essential for plant growth, development, and environmental adaptions, only few reports exist on plant telomeric G4 DNA (pTG4). Here, using bulk and single-molecule assays, including CD spectroscopy, and single-molecule FRET approaches, we comprehensively characterized the structure and dynamics of a typical plant telomeric sequence, d[GGG(TTTAGGG)3]. We found that this sequence can fold into mixed G4s in potassium, including parallel and antiparallel structures. We also directly detected intermediate dynamic transitions, including G-hairpin, parallel G-triplex, and antiparallel G-triplex structures. Moreover, we observed that pTG4 is unfolded by the AtRecQ2 helicase but not by AtRecQ3. The results of our work shed light on our understanding about the existence, topological structures, stability, intermediates, unwinding, and functions of pTG4.

Intestinal schistosomiasis masquerading as intestinal polyps.

BACKGROUND: Schistosomiasis is very common in the southern part of the Yangtze River Basin in China. It is mainly manifested as appendicitis, ulcers, hematomas, and thickening of the intestinal tract. Schistosomiasis of the appendix is rare, mainly manifested as appendicitis, which is easy to be misdiagnosed. CASE PRESENTATION: Here we report a rare case of a Chinese female whose intestinal mass manifested as intestinal polyps and was eventually diagnosed pathologically as schistosomiasis infection (appendix schistosomiasis). So far, there are rare relevant cases reported. CONCLUSIONS: Intestinal schistosomiasis is easily misdiagnosed, and appendix schistosomiasis is rare. The final diagnosis requires pathology, especially surgical pathology.

Studying the Potassium-Induced G-Quadruplex DNA Folding Process Using Microscale Thermophoresis.

Guanine (G) quadruplexes (G4s) can be formed by G-rich sequences when stabilized by the binding of cations (typically K+ or Na+) and play an essential role in replication, recombination, transcription, and telomere maintenance. Understanding of the G4 folding process is crucial for determining their cellular functions. However, G4-K+ interactions and folding pathways are still not well understood. By using human telomeric G4 (hTG4) as an example, two binding states corresponding to two K+ cations binding to hTG4 were distinguished clearly and fitted precisely. The basic binding parameters during G4-K+ interactions were measured and calculated by taking advantage of microscale thermophoresis (MST), which monitors the changes in charge and size at the same time. The G-hairpin and G-triplex have been suggested as intermediates during G4 folding and unfolding. We further analyzed the equilibrium dissociation constants of 10 possible folding intermediates using MST; thus, the energetically favorable folding/unfolding pathways were proposed. The results might not only shed new light on G4-K+ interactions and G4 folding pathways but also provide an example for experimentally studying DNA-ion interactions.

A comprehensive analysis of the allergenicity and IgE epitopes of myosinogen allergens in Scylla paramamosain.

BACKGROUND: Scylla paramamosain is one of the most common and serious food allergens in Asia. Therefore, research on its prevalence, accurate diagnosis, and IgE-binding pattern of the allergens is crucial. OBJECTIVE: To identify the IgE epitopes of the myosinogen allergens in S. paramamosain using phage peptide library. METHODS: The prevalence of allergy to crabs (AC) and of sensitization was analysed using a questionnaire and a serological assay. BAT was performed by flow cytometry, and its diagnostic performance was evaluated in relation to allergens purified from crab myosinogen. IgE-binding epitopes were identified by phage display using the IgE from patients with AC. Sequence- and structure-based bioinformatics analyses were performed to identify allergenic epitopes. RESULTS: Crab was the most common cause of food allergies in this study. Subjects with AC (n = 30) with clear clinical symptoms were identified by immunoblotting and BAT. All of the myosinogen allergens triggered basophil activation; surface expression of CD63 and CD203c was higher in patients allergic to AK and FLN c than in patients allergic to SCP and TIM. In addition to six conformational epitopes of SCP, six linear epitopes and eight conformational epitopes of AK were identified. Five linear epitopes and three conformational epitopes of TIM, nine linear and ten conformational epitopes of FLN c were also identified, and the sequence VH(I/T) L was appeared in epitopes of both TIM and FLN c. The number of epitopes showed consistency with the value of BAT. CONCLUSIONS AND CLINICAL RELEVANCE: BAT can be used for accurate diagnosis of AC. Identification of particular allergenic motifs could be a valuable tool for prevention, diagnosis, and treatment of food allergies.

Phylogeography of the arid shrub Atraphaxis frutescens (Polygonaceae) in northwestern China: evidence from cpDNA sequences.

Climatic fluctuations during the Pleistocene are usually considered as a significant factor in shaping intraspecific genetic variation and influencing demographic histories. To well-understand these processes in desert northwest China, we selected arid adapted Atraphaxis frutescens as the study species. Two cpDNA regions (psbK-psbI, psbB-psbH) were sequenced in 272 individuals from 33 natural populations across the range of this shrub, and 10 haplotypes were identified. It was found to contain high levels of total gene diversity (H T = 0.858), and low levels of within-population diversity (H S = 0.092). Analysis of molecular variance (AMOVA) indicates that genetic differentiation primarily occurs among groups of populations. Based on BEAST (Bayesian Evolutionary Analysis Sampling Trees) analysis, we suggest that intraspecific differentiation of the species, resulting from isolated populations, accompanied enhanced desertification during the middle and late Pleistocene. The expansion of the Gurbantunggut and Kumtag deserts in this area appears to have triggered divergence among populations of the western, central, and eastern portions of the region and shaped genetic differentiation among them. Two possible independent glacial refugia were predicted, the Ili Valley and the northern Junggar Basin. Extensive development of arid habitats (desert margin and arid piedmont grassland) coupled with a more equable climate because the early Holocene are factors likely to have generated recent expansion of A. frutescens.

Phylogeographical structure inferred from cpDNA sequence variation of Zygophyllum xanthoxylon across north-west China.

Zygophyllum xanthoxylon, a desert species, displaying a broad east-west continuous distribution pattern in arid Northwestern China, can be considered as a model species to investigate the biogeographical history of this region. We sequenced two chloroplast DNA spacers (psbK-psbI and rpl32-trnL) in 226 individuals from 31 populations to explore the phylogeographical structure. Median-joining network was constructed and analysis of AMOVA, SMOVA, neutrality tests and distribution analysis were used to examine genetic structure and potential range expansion. Using species distribution modeling, the geographical distribution of Z. xanthoxylon was modeled during the present and at the Last Glacial Maximum (LGM). Among 26 haplotypes, one was widely distributed, but most was restricted to either the eastern or western region. The populations with the highest levels of haplotype diversity were found in the Tianshan Mountains and its surroundings in the west, and the Helan Mountains and Alxa Plateau in the east. AMOVA and SAMOVA showed that over all populations, the species lacks phylogeographical structure, which is speculated to be the result of its specific biology. Neutrality tests and mismatch distribution analysis support past range expansions of the species. Comparing the current distribution to those cold and dry conditions in LGM, Z. xanthoxylon had a shrunken and more fragmented range during LGM. Based on the evidences from phylogeographical patterns, distribution of genetic variability, and paleodistribution modeling, Z. xanthoxylon is speculated most likely to have originated from the east and migrated westward via the Hexi Corridor.

Tertiary montane origin of the Central Asian flora, evidence inferred from cpDNA sequences of Atraphaxis (Polygonaceae).

Atraphaxis has approximately 25 species and a distribution center in Central Asia. It has been previously used to hypothesize an origin from montane forest. We sampled 18 species covering three sections within the genus and sequenced five cpDNA spacers, atpB-rbcL, psbK-psbI, psbA-trnH, rbcL, and trnL-trnF. BEAST was used to reconstruct phylogenetic relationship and time divergences, and S-DIVA and Lagrange were used, based on distribution area and ecotype data, for reconstruction of ancestral areas and events. Our results appear compatible with designation of three taxonomic sections within the genus. The generic stem and crown ages were Eocene, approximately 47 Ma, and Oligocene 27 Ma, respectively. The origin of Atraphaxis is confirmed as montane, with an ancestral area consisting of the Junggar Basin and uplands of the Pamir-Tianshan-Alatau-Altai mountain chains, and ancestral ecotype of montane forest. Two remarkable paleogeographic events, shrinkage of the inland Paratethys Sea at the boundary of the late Oligocene and early Miocene, and the time intervals of cooling and drying of global climate from 24 (22) Ma onward likely facilitated early diversification of Atraphaxis, while rapid uplift of the Tianshan Mountains during the late Miocene may have promoted later diversification.

Interplay Between Intracellular Transport Dynamics and Liquid‒Liquid Phase Separation.

Liquid‒liquid phase separation (LLPS) is a ubiquitous process in which proteins, RNA, and biomolecules assemble into membrane-less compartments, playing important roles in many biological functions and diseases. The current knowledge on the biophysical and biochemical principles of LLPS is largely from in vitro studies; however, the physiological environment in living cells is complex and not at equilibrium. The characteristics of intracellular dynamics and their roles in physiological LLPS remain to be resolved. Here, by using single-particle tracking of quantum dots and dynamic monitoring of the formation of stress granules (SGs) in single cells, the spatiotemporal dynamics of intracellular transport in cells undergoing LLPS are quantified. It is shown that intracellular diffusion and active transport are both reduced. Furthermore, the formation of SG droplets contributes to increased spatial heterogeneity within the cell. More importantly, the study demonstrated that the LLPS of SGs can be regulated by intracellular dynamics in two stages: the reduced intracellular diffusion promotes SG assembly and the microtubule-associated transport facilitates SG coalescences. The work on intracellular dynamics not only improves the understanding of the mechanism of physiology phase separations occurring in nonequilibrium environments but also reveals an interplay between intracellular dynamics and LLPS.

Diversification of plant species in arid Northwest China: species-level phylogeographical history of Lagochilus Bunge ex Bentham (Lamiaceae).

Lagochilus occurs in the arid zones across temperate steppe and desert regions of Northwest China. Cooling with strong dessication in the Pleistocene, along with rapid uplift of mountain ranges peripheral to the Qinghai-Tibet Plateau, appear to have had major impacts on the genetic structure of the flora. To understand the evolutionary history of Lagochilus and the divergence related to these past shifts of habitats among these regions, we sequenced the plastid intergenic spacers, psbA-trnH and trnS-trnG from populations throughout the known distributions of ten species of the genus. We investigated species-level phylogeographical patterns within Lagochilus. Phylogenetic trees were constructed using Neighbor-joining and Bayesian inference. The divergence times of major lineages were estimated with BEAST and IMa. Genetic structure and demographic history were inferred by AMOVA, neutrality tests, mismatch distribution, and Bayesian skyline plot analyses. The results showed that most chloroplast haplotypes were species-specific, and that the phylogeny of Lagochilus is geographically structured. The estimated Bayesian chronology and IMa suggested that the main divergence events for species between major eastern and western portions of the Chinese desert occurred at the Plio-/Pleistocene boundary (ca. 2.1-2.8 Ma ago), and likely coinciding with the formation of these deserts in Northwest China. The regional demographic expansions, in the western region at ca. 0.39 Ma, and in the eastern at ca. 0.06 Ma, or across all regions at ca. 0.26 Ma, showed the response to aridification accompanied by cooling of the Pleistocene sharply increased aridity in the Chinese deserts, which reflects a major influence of geologic and climatic events on the evolution of species of Lagochilus. We suggest that diversification is most likely the result of the past fragmentation due to aridification; the expansion of the range of species along with the deserts was an adaptation to dry and cold environments during the Quaternary.

Intracellular transport dynamics revealed by single-particle tracking.

Intracellular transport is the basis for the transfer of matter, energy, and information in cells and is critical to many cellular functions. Within the nonequilibrium environment of living cells, the transport behaviours are far from the traditional motion in liquid but are more complex and active. With the advantage of high spatial and temporal resolution, the single-particle tracking (SPT) method is widely utilized and has achieved great advances in revealing intracellular transport dynamics. This review describes intracellular transport from a physical perspective and classifies it into two modes: diffusive motion and directed motion. The biological functions and physical mechanisms for these two transport modes are introduced. Next, we review the principle of SPT and its advances in two aspects of intracellular transport. Finally, we discuss the prospect of near infrared SPT in exploring the in vivo intracellular transport dynamics.

miRNA-186 Improves Sepsis Induced Renal Injury Via PTEN/PI3K/AKT/P53 Pathway.

AIM: The aim of this study is to explain the effects of miRNA-186 in renal injury induced by sepsis. METHODS: The Wistar rats were divided into 3 groups: the Sham group, Sepsis model group and the miRNA-186 group based on the model group; there were 9 rats in every group. The rat sepsis model was reproduced by cecal ligation and puncture (CLP). The rats of the miRNA-186 group were injected miRNA-186 from caudal vein. The rats of the difference group were killed after operation 24 h. The kidneys of the difference groups were taken for histopathological and cell apoptosis analysis by H&E and TUNEL assay. The relative protein expressions were measured by WB assay. miRNA-186 target to Phosphatase and tensin homologous protein (PTEN). RESULTS: Compared with the Sham group, the kidney histopathological and cell apoptosis rates of the model group were significantly damaged (P<0.05, respectively), however, the kidney histopathological and cell apoptosis rate of miRNA-186 group were significantly improved compared with the model group (P<0.05, respectively). The relative protein expressions were significantly different among 3 groups (P<0.05, respectively). The PTEN was the target of the miRNA-186. CONCLUSION: miRNA-186 over-expression has effects that improve renal injury induced by sepsis via PTEN pathway.

Expression and epitope identification of myosin light chain isoform 1, an allergen in Procambarus clarkii.

Myosin light chain isoform 1 (MLC1) is reported to be a novel allergen in crayfish (Procambarus clarkii). However, little information is available about its allergic epitopes. In this study, recombinant crayfish MLC1 (rMLC1) was expressed and confirmed by mass spectrometry. Circular dichroic analysis and serological test were performed for the measuring of structural and immunological properties of rMLC1. Specific-protein-A-enriched IgG raised in rabbits against purified rMLC1 was used to screen a phage display random peptide library. Nine MLC1 mimotope clones were identified among 16 random clones after biopanning. Five conformational epitopes were identified with the program LocaPep, and mapped into 3 epitope regions at the antibody-binding interface of MLC1. MLC1 of crayfish showed high primary and secondary structure identity to MLC of other allergenic species, its epitopes were located in the structure conserved regions, and its cross-reactivity among related species was indicated by immunological assays.

[Investigations of influential factors of acute renal injury after coronary artery bypass grafting].

OBJECTIVE: To investigate the incidence, correlate factors and effect on convalescent of acute kidney injury (AKI) after coronary artery bypass grafting (CABG). METHODS: Sixty-eight consecutive patients who underwent scheduled CABG were included. AKI was diagnosed according to the diagnostic criteria from expert group of AKI. The patients were divided into AKI group with AKI and control group without AKI. Difference of clinic features was contrasted between the two groups. And multivariable logistic regression modeling was used to identify the risk factors of AKI. RESULTS: AKI occurred in 27.94% patients (19/68). There was significant difference between the two groups in gender, hypertension, hyperlipemia, number of grafts, the duration of mechanic ventilation, the time of ICU and the length of stay post operation. Hypertension(Wald 3.27, P=0.07, 95% CI 0.85-53.59), number of grafts(Wald 7.67, P=0.01, 95%CI 1.48-9.73), the duration of mechanic ventilation(Wald 4.94, P=0.03, 95%CI 1.00-1.18) were risk factors of AKI in multivariable logistic regression modeling. CONCLUSION: The incidence of AKI was high (27.94%). Female, hypertension, hyperlipemia, number of grafts, the duration of mechanic ventilation were risk factors in monovariance analysis, and AKI may increase the time of ICU and the length of stay post operation. Hypertension, number of grafts, duration of mechanic ventilation were independent risk factors of AKI in multivariable logistic regression model.

Pleistocene climate change and phylogeographic structure of the Gymnocarpos przewalskii (Caryophyllaceae) in the northwest China: Evidence from plastid DNA, ITS sequences, and Microsatellite.

Northwestern China has a wealth of endemic species, which has been hypothesized to be affected by the complex paleoclimatic and paleogeographic history during Quaternary. In this paper, we used Gymnocarpos przewalskii as a model to address the evolutionary history and current population genetic structure of species in northwestern China. We employed two chloroplast DNA fragments (rps16 and psbB-psbI), one nuclear DNA fragment (ITS), and simple sequence repeat (SSRs) to investigate the spatial genetic pattern of G. przewalskii. High genetic diversity (cpDNA: h S = 0.330, h T = 0.866; ITS: h S = 0.458, h T = 0.872) was identified in almost all populations, and most of the population have private haplotypes. Moreover, multimodal mismatch distributions were observed and estimates of Tajima's D and Fu's FS tests did not identify significantly departures from neutrality, indicating that recent expansion of G. przewalskii was rejected. Thus, we inferred that G. przewalskii survived generally in northwestern China during the Pleistocene. All data together support the genotypes of G. przewalskii into three groups, consistent with their respective geographical distributions in the western regions-Tarim Basin, the central regions-Hami Basin and Hexi Corridor, and the eastern regions-Alxa Desert and Wulate Prairie. Divergence among most lineages of G. przewalskii occurred in the Pleistocene, and the range of potential distributions is associated with glacial cycles. We concluded that climate oscillation during Pleistocene significantly affected the distribution of the species.

Reducing Allergenicity to Arginine Kinase from Mud Crab Using Site-Directed Mutagenesis and Peptide Aptamers.

The mud crab ( Scylla paramamosain) is widely consumed but can cause a severe food allergic reaction. To reduce allergenicity to arginine kinase (AK), site-directed mutagenesis was used to destroy disulfide bonds or mutate critical amino acids of conformational epitopes. Three hypoallergenic mutant AKs (mAK1, mAK2, and mAK3) were generated, with the immunoreactivity decreasing by 54.2, 40.1, and 71.4%, respectively. In comparison to recombinant AK (rAK), the structure of mAKs was clearly changed. Additionally, antisense peptides were designed on the basis of linear epitopes and pepsin-cutting sites of AK. Five peptide aptamers were screened by molecular docking and then analyzed by the immunoglobulin E inhibition enzyme-linked immunosorbent assay and human Laboratory of Allergic Diseases 2 mast cell degranulation assay. The peptide aptamers could significantly inhibit allergenicity of rAK and mAKs, and the inhibitory effect of peptide aptamer 3 was slightly better than the others. These results provide synergistic methods to reduce allergenicity to AK, which could be applied to other shellfish allergens.

Analysis of the Allergenic Epitopes of Tropomyosin from Mud Crab Using Phage Display and Site-Directed Mutagenesis.

Mud crab ( Scylla serrata), which is widely consumed, can cause severe allergic symptoms. Eight linear epitopes and seven conformational epitopes of tropomyosin (TM) from S. serrata were identified using phage display. The conformational epitopes were formed based on the coiled-coil structure of TM. Most of the epitopes were located in the regions where primary structures were conserved among crustacean TM. Twelve synthetic peptides were designed according to the epitopes and trypsin-cutting sites of TM, among them, three synthetic peptides (including one linear epitope and two conformational epitopes) were recognized by all of the patient sera using inhibitory dot blotting. A triple-variant (R90A-E164A-Y267A) was constructed based on the critical amino acids of the TM epitope. The IgE-binding activity of the triple-variant was significantly reduced compared with that of native TM. The results of phage display and site-directed mutagenesis offered new information regarding conformational epitopes of TM.

Risk factors for mortality after coronary artery bypass grafting in patients with low left ventricular ejection fraction.

BACKGROUND: We managed to assess and confirm the risk factors for mortality after coronary artery bypass grafting (CABG) operations so as to map out the proper guidance of surgical strategy especially in patients with low left ventricular ejection fraction (LVEF) in domestic polyclinic like ours. METHODS: Five hundred and forty-eight consecutive patients underwent CABG from December 1999 through August 2005 were analyzed retrospectively. Eighty-nine cases had an LVEF of 40% or less. All together twenty-two candidate factors were evaluated for their association with perioperative death using univariate and multivariate stepwise Logistic analysis. RESULTS: When data from all the patients who had undergone CABG were taken into account, LVEF, left ventricular end diastolic diameter (LVEDD), mitral regurgitation, aneurysm of the heart wall, mitral repair/replacement, resection of aneurysm, concomitant aortic valve replacement, and perioperative intra-aortic balloon counter-pulsation (IABP), left ventricular assist device (LVAD) and cardiopulmonary bypass (CPB) all showed an association with perioperative death in univariate analysis, while an LVEF of > 40%, on the other hand, appeared to be a protective factor. In multivariate analysis, moderate to severe mitral regurgitation, aneurysm of the heart wall, repair of septal perforation and aortic regurgitation were proved to be risk factors. When the analysis was restricted to patients with an LVEF of 40% or less, such variables as age, LVEDD, mitral regurgitation, mitral repair/replacement, IABP, and CPB were qualified as risk factors in a univariate analysis. Age, moderate mitral regurgitation, aneurysm of the heart wall, CPB, left main coronary artery disease and female were associated with perioperative death in a multivariate logistic regression analysis. CONCLUSIONS: Concerning the prognosis, patients who undergo CABG would have different risk factors when data from all the enrolled patients or data from patients with LVEF

[Treatment of a case of Kawasaki's disease complicated with myocardial infarction].

Coronary artery by-pass graft (CABG) and ventricular aneurysmectomy were done for a 2.5 years old girl with Kawasaki's disease complicated by myocardial ischemia and old myocardial infarction, left ventricular aneurysm formation and heart failure. Post-operatively, the cardiac out-put of the patient increased, the heart failure syndrome disappeared, and cardiac function improved to class I(NYHA) one month later. In the 6-year follow-up, the patient kept her heart function at class I, and had normal growth and development. Coronary artery by-pass graft is an effective therapeutic strategy for childhood myocardial ischemia, and attention should be paid to some special considerations.

Spatial Patterns of Species Diversity and Phylogenetic Structure of Plant Communities in the Tianshan Mountains, Arid Central Asia.

The Tianshan Mountains, located in arid Central Asia, have a humid climate and are biodiversity hotspots. Here, we aimed to clarify whether the pattern of species diversity and the phylogenetic structure of plant communities is affected by environmental variables and glacial refugia. In this study, plant community assemblies of 17 research sites with a total of 35 sample plots were investigated at the grassland/woodland boundaries on the Tianshan Mountains. Community phylogeny of these plant communities was constructed based on two plant DNA barcode regions. The indices of phylogenetic diversity and phylogenetic community structure were calculated for these sample plots. We first estimated the correlation coefficients between species richness (SR) and environmental variables as well as the presence of glacial refugia. We then mapped the significant values of indices of community phylogeny (PD, RPD, NRI, and NTI) to investigate the correlation between community phylogeny and environmental structure or macrozones in the study area. The results showed that a significantly higher value of SR was obtained for the refugial groups than for the colonizing groups (P < 0.05); presence of refugia and environmental variables were highly correlated to the pattern of variation in SR. Indices of community phylogeny were not significantly different between refugial and colonizing regions. Comparison with the humid western part showed that plant communities in the arid eastern part of the Tianshan Mountains tended to display more significant phylogenetic overdispersion. The variation tendency of the PhyloSor index showed that the increase in macro-geographical and environmental distance did not influence obvious phylogenetic dissimilarities between different sample plots. In conclusion, glacial refugia and environmental factors profoundly influenced the pattern of SR, but community phylogenetic structure was not affected by glacial refugia among different plant communities on the Tianshan Mountains. This pattern of community phylogenetic structure could have resulted from shared ancestry and species pool among these sample plots.

Insight into Central Asian flora from the Cenozoic Tianshan montane origin and radiation of Lagochilus (Lamiaceae).

The Tianshan Mountains play a significant role in the Central Asian flora and vegetation. Lagochilus has a distribution concentration in Tianshan Mountains and Central Asia. To investigate generic spatiotemporal evolution, we sampled most Lagochilus species and sequenced six cpDNA locations (rps16, psbA-trnH, matK, trnL-trnF, psbB-psbH, psbK-psbI). We employed BEAST Bayesian inference for dating, and S-DIVA, DEC, and BBM for ancestral area/biome reconstruction. Our results clearly show that the Tianshan Mountains, especially the western Ili-Kirghizia Tianshan, as well as Sunggar and Kaschgar, was the ancestral area. Ancestral biome was mainly in the montane steppe zone of valley and slope at altitudes of 1700-2700 m, and the montane desert zone of foothill and front-hill at 1000-1700 m. Here two sections Inermes and Lagochilus of the genus displayed "uphill" and "downhill" speciation process during middle and later Miocene. The origin and diversification of the genus were explained as coupled with the rapid uplift of the Tianshan Mountains starting in late Oligocene and early Miocene ca. 23.66~19.33 Ma, as well as with uplift of the Qinghai-Tibetan Plateau (QTP) and Central Asian aridification.