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Monoallelic VSG expression is essential for Trypanosoma brucei survival. Competition between TbRAP1’s RNA and dsDNA binding activities ensures that TbRAP1 sustains a high level expression of the active VSG while silencing other VSGs globally.

The aim of this study was to investigate if the supplementation of folic acid and taurine can relieve the adverse effects of different levels of heat stress (HS) on growth performance, physiological indices, antioxidative capacity, immunity, rumen fermentation and microbiota. A total of 24 Dorper × Hu crossbred lambs (27.51 ± 0.96 kg) were divided into four groups: control group (C, 25 °C), moderate HS group (MHS, 35 °C), severe HS group (SHS, 40 °C), and the treatment group, under severe HS (RHS, 40 °C, 4 and 40 mg/kg BW/d coated folic acid and taurine, respectively). Results showed that, compared with Group C, HS significantly decreased the ADG of lambs (p < 0.05), and the ADG in the RHS group was markedly higher than in the MHS and SHS group (p < 0.05). HS had significant detrimental effects on physiological indices, antioxidative indices and immune status on the 4th day (p < 0.05). The physiological indices, such as RR and ST, increased significantly (p < 0.05) with the HS level and were significantly decreased in the RHS group, compared to the SHS group (p < 0.05). HS induced the significant increase of MDA, TNF-α, and IL-β, and the decrease of T-AOC, SOD, GPx, IL-10, IL-13, IgA, IgG, and IgM (p < 0.05). However, there was a significant improvement in these indices after the supplementation of folic acid and taurine under HS. Moreover, there were a significant increase in Quinella and Succinivibrio, and an evident decrease of the genera Rikenellaceae_RC9_gut_group and Asteroleplasma under HS (p < 0.05). The LEfSe analysis showed that the genera Butyrivibrio, Eubacterium_ventriosum_group, and f_Bifidobacteriaceae were enriched in the MHS, SHS and RHS groups, respectively. Correlated analysis indicated that the genus Rikenellaceae_RC9_gut_group was positively associated with MDA, while it was negatively involved in IL-10, IgA, IgM, and SOD (p < 0.05); The genus Anaeroplasma was positively associated with the propionate and valerate, while the genus Succinivibrio was negatively involved in TNF-α (p < 0.05). In conclusion, folic acid and taurine may alleviate the adverse effects of HS on antioxidant capacity, immunomodulation, and rumen fermentation of lambs by inducing changes in the microbiome that improve animal growth performance.

Telomere maintenance is essential for genome integrity and chromosome stability in eukaryotic cells harboring linear chromosomes, as telomere forms a specialized structure to mask the natural chromosome ends from DNA damage repair machineries and to prevent nucleolytic degradation of the telomeric DNA. In Trypanosoma brucei and several other microbial pathogens, virulence genes involved in antigenic variation, a key pathogenesis mechanism essential for host immune evasion and long-term infections, are located at subtelomeres, and expression and switching of these major surface antigens are regulated by telomere proteins and the telomere structure. Therefore, understanding telomere maintenance mechanisms and how these pathogens achieve a balance between stability and plasticity at telomere/subtelomere will help develop better means to eradicate human diseases caused by these pathogens. Telomere replication faces several challenges, and the “end replication problem” is a key obstacle that can cause progressive telomere shortening in proliferating cells. To overcome this challenge, most eukaryotes use telomerase to extend the G-rich telomere strand. In addition, a number of telomere proteins use sophisticated mechanisms to coordinate the telomerase-mediated de novo telomere G-strand synthesis and the telomere C-strand fill-in, which has been extensively studied in mammalian cells. However, we recently discovered that trypanosomes lack many telomere proteins identified in its mammalian host that are critical for telomere end processing. Rather, T. brucei uses a unique DNA polymerase, PolIE that belongs to the DNA polymerase A family (E. coli DNA PolI family), to coordinate the telomere G- and C-strand syntheses. In this review, I will first briefly summarize current understanding of telomere end processing in mammals. Subsequently, I will describe PolIE-mediated coordination of telomere G- and C-strand synthesis in T. brucei and implication of this recent discovery.

Recently, the relationship between the goat host and its gastrointestinal microbiome has emerged as a hallmark of host-microbiota symbiosis, which was indispensable for the proper physiological function that convert the plant biomass to livestock products. However, little integrative information about the establishment of gastrointestinal microflora in goats exists. Herein, we characterized the colonizing process of the bacterial community in the digesta and mucosa of the rumen, cecum, and colon of the cashmere goat from birth to adulthood to compare its spatiotemporal difference via 16S rRNA gene sequencing. A total of 1,003 genera belonging to 43 phyla were identified. Principal coordinate analysis unveiled the similarity of microbial community between or within each age group increased and gradually developed toward mature whatever in digesta or mucosa. In the rumen, the composition of the bacterial community in digesta differed significantly from in mucosa across age groups; whereas in the hindgut, there was a high similarity of bacterial composition between the in digesta and mucosa in each age group before weaning, while the bacterial community structure differed markedly between these two types of samples after weaning. Taxonomic analysis indicated that 25 and 21 core genera coexisted in digesta and mucosa of the rumen and hindgut, respectively; but their abundances differed considerably by GIT region and/or age. In digesta, as goats aged, a lower abundance of Bacillus was observed with higher abundances of Prevotella 1 and Rikenellaceae RC9 in the rumen; while in the hindgut, the genera Escherichia-Shigella, Variovorax, and Stenotrophomonas decreased and Ruminococcaceae UCG-005, Ruminococcaceae UCG-010, and Alistipes increased with age increased. In mucosa, the rumen showed microbial dynamics with increases of Butyrivibrio 2 and Prevotellaceae UCG-001 and decreases of unclassified_f_Pasteurellaceae; while the genera Treponema 2 and Ruminococcaceae UCG-010 increased and Escherichia-Shigella decreased in the hindgut as goats aged. These results shed light on the colonization process of microbiota in the rumen and hindgut, which mainly include the initial, transit, and mature phases. Furthermore, there is a significant difference in the microbial composition between in digesta and mucosa, and both these exhibit a considerable spatiotemporal specificity.

Although located at the chromosome end, telomeres are an essential chromosome component that helps maintain genome integrity and chromosome stability from protozoa to mammals. The role of telomere proteins in chromosome end protection is conserved, where they suppress various DNA damage response machineries and block nucleolytic degradation of the natural chromosome ends, although the detailed underlying mechanisms are not identical. In addition, the specialized telomere structure exerts a repressive epigenetic effect on expression of genes located at subtelomeres in a number of eukaryotic organisms. This so-called telomeric silencing also affects virulence of a number of microbial pathogens that undergo antigenic variation/phenotypic switching. Telomere proteins, particularly the RAP1 homologs, have been shown to be a key player for telomeric silencing. RAP1 homologs also suppress the expression of Telomere Repeat-containing RNA (TERRA), which is linked to their roles in telomere stability maintenance. The functions of RAP1s in suppressing telomere recombination are largely conserved from kinetoplastids to mammals. However, the underlying mechanisms of RAP1-mediated telomeric silencing have many species-specific features. In this review, I will focus on Trypanosoma brucei RAP1’s functions in suppressing telomeric/subtelomeric DNA recombination and in the regulation of monoallelic expression of subtelomere-located major surface antigen genes. Common and unique mechanisms will be compared among RAP1 homologs, and their implications will be discussed.

Telomerase is a ribonucleoprotein enzyme responsible for maintaining the telomeric end of the chromosome. The telomerase enzyme requires two main components to function: the telomerase reverse transcriptase (TERT) and the telomerase RNA (TR), which provides the template for telomeric DNA synthesis. TR is a long non-coding RNA, which forms the basis of a large structural scaffold upon which many accessory proteins can bind and form the complete telomerase holoenzyme. These accessory protein interactions are required for telomerase activity and regulation inside cells. The interacting partners of TERT have been well studied in yeast, human, and Tetrahymena models, but not in parasitic protozoa, including clinically relevant human parasites. Here, using the protozoan parasite, Trypanosoma brucei (T. brucei) as a model, we have identified the interactome of T. brucei TERT (TbTERT) using a mass spectrometry-based approach. We identified previously known and unknown interacting factors of TbTERT, highlighting unique features of T. brucei telomerase biology. These unique interactions with TbTERT, suggest mechanistic differences in telomere maintenance between T. brucei and other eukaryotes.

Background: Unlocking phenotype plasticity (UPP) has been shown to have an essential role in the mechanism of tumor development and therapeutic response. However, the clinical significance of unlocking phenotypic plasticity in patients with lung adenocarcinoma is unclear. This study aimed to explore the roles of unlocking phenotypic plasticity in immune status, prognosis, and treatment in patients with lung adenocarcinoma (LUAD).Methods: Differentially expressed genes (DEGs) and clinical information of UPP were selected from the cancer genome atlas (TCGA) database, and the GO, KEGG enrichment analyses were performed. The independent prognostic genes were determined by univariate and multivariate Cox regression, and the UPP signature score was constructed. Patients with LUAD were divided into high- and low-risk groups according to the median of score, and the immunocytes and immune function, the gene mutation, and drug sensitivities between the two groups were analyzed. Finally, the results were validated in the GEO database.Results: Thirty-nine significantly DEGs were determined. Enrichment analysis showed that UPP-related genes were related to protein polysaccharides and drug resistance. The prognostic results showed that the survival of patients in the high-risk group was poorer than that in the low-risk group (p < 0.001). In the high- and low-risk groups, single nucleotide polymorphism (SNP) and C > T are the most common dissent mutations. The contents of immune cells were significantly different between high- and low-risk groups. And the immune functions were also significantly different, indicating that UPP affects the immunity in LUAD. The results from TCGA were validated in the GEO.Conclusion: Our research has proposed a new and reliable prognosis indicator to predict the overall survival. Evaluation of the UPP could help the clinician to predict therapeutic responses and make individualized treatment plans in patients with LUAD.

Abstract Background Preweaned rumen development is vital for animal health and efficient fermentation. In this study, we integrated ruminal transcriptomic and metagenomic data to explore the dynamics of rumen functions, microbial colonization, and their functional interactions during the first 8 weeks of life in goats. Results The dynamic rumen transcriptomic and microbial profiles both exhibited two distinct phases during early rumen development. The differentially expressed genes of the rumen transcriptome between the two phases showed that the immune-related response was enriched in the first phase and nutrient-related metabolism was enriched in the second phase, whereas the differentially expressed genes of the rumen microbiome were enriched in bacteriocin biosynthesis and glycolysis/gluconeogenesis activities. The developmental shift in the rumen transcriptome (at d 21) was earlier than the feed stimulus (at d 25) and the shift in the rumen microbiome (at d 42). Additionally, 15 temporal dynamic rumen gene modules and 20 microbial modules were revealed by coexpression network analysis. Functional correlations between the rumen and its microbiome were primarily involved in rumen pH homeostasis, nitrogen metabolism and the immune response. Rumen gene modules associated with the microbial alpha diversity index were also enriched in the immune response process. Conclusions The present study touched the critical developmental process of rumen functions, microbial colonization and their functional interactions during preweaned development. Taken together, these results demonstrated that rumen development at the first phase is more likely a programmed process rather than stimulation from feed and the microbiome, while the shift of rumen metagenomes was likely regulated by both the diet and host. The intensive functional correlations between rumen genes and the microbiome demonstrated that synergistic processes occurred between them during early rumen development.

In this paper, the effect of batch-annealing temperature on the oxidation of 22MnB5 steel during austenitizing was studied. When the batch-annealing temperature was decreased from 690 to 680 °C, the surface roughness of cold-rolled 22MnB5 steel decreased, which reduced the oxide layer thickness during the subsequent austenitizing process and had little effect on the mechanical properties in the cold-rolled and quenched state. This indicated that oxidation during austenitizing could be reduced by properly reducing the batch-annealing temperature without affecting the mechanical properties.

ABSTRACT: Background: Thulium laser (Tm:YAG) prostate surgery is a safe and effective procedure with low morbidity and comparable clinical outcomes to those of transurethral resection of the prostate (TURP). However, the sexual function outcomes (erectile and ejaculatory function) have been scarcely studied. Aim: We aimed to assess the impact of Tm:YAG prostate surgery on sexual outcomes (erectile and ejaculatory function) and compare them with those patients undergoing TURP. Material and Methods: We searched digital databases like PUBMED, SCOPUS, CENTRAL and EMBASE using relevant keywords to identify comparative studies on TURP and non-comparative studies on Tm:YAG prostate surgery that assessed sexual outcomes. We performed qualitative and quantitative analyses with the extracted data. We carried out a meta-analysis to compare postoperative International Index of Erectile Function (IIEF-5) scores and incidences of retrograde ejaculation (RE) in patients undergoing either Tm:YAG or TURP. The pre-operative and post-operative IIEF-5 scores were pooled to estimate overall scores. Results: We included 5 comparative and 8 non-comparative studies in this review. We found the postoperative IIEF-5 score improvements to be significantly higher in the Tm:YAG prostate surgery group than in the TURP group with a significant mean difference (MD) of 0.45 (95% CI, 0.18 to 0.72; P = .001). We found no significant associations between the procedures. The pooled OR for the association of RE was estimated at 0.90 (95% CI, 0.50 to 1.60; P = .71; I2 = 0%). Conclusion: Tm:YAG prostate surgery improves erectile function more than TURP, according to our findings. Tm:YAG prostate aided surgery also outperforms TURP in terms of preserving sexual function following surgery.However, We found similar or no difference in incidence of RE between Tm:YAG prostate surgery and TURP.Bibo L, Hao L, Pang K, et al. Assessment of Sexual Outcomes in Patients Undergoing Thulium Laser Prostate Surgery for Management of Benign Prostate Hyperplasia: A Systematic Review and Meta-analysis. Sex Med 2022;10:100483.

The intestine is a complex micro-ecosystem, and its stability determines the health of animals. Different dietary nutritional levels affect the intestinal microbiota and health. In this study, the nutritional levels of energy and protein in the diet of goats were changed, and the body weight was measured every 15 days. In the late feeding period, 16 S rRNA sequencing technology was used to detect the content of microorganisms. A meteorological chromatograph was used to detect volatile fatty acids in the cecum and colon of goats. In the feeding stage, reducing the nutritional level of the diet significantly reduced the weight of the lamb (p < 0.05). In the cecum, the abundance of potentially harmful bacteria, such as Sphingomonas, Marvinbryantia, and Eisenbergiella, were significantly enriched in goats fed with the standard nutritional level diets (p < 0.05). Additionally, the contents of acetate (p = 0.037) and total VFAs (p = 0.041) increased. In the colon, the abundance of SCFAs-producing bacteria, such as Ruminococcaceae, Christensenellaceae, and Papillibacter, decreased as the nutritional level in the diet increased (p < 0.05). In conclusion, the increase in nutritional level could affect the growth performance and composition of intestinal microbiota.

Telomeres, the nucleoprotein complexes at chromosome ends, are well-known for their essential roles in genome integrity and chromosome stability. Yet, telomeres and subtelomeres are frequently less stable than chromosome internal regions. Many subtelomeric genes are important for responding to environmental cues, and subtelomeric instability can facilitate organismal adaptation to extracellular changes, which is a common theme in a number of microbial pathogens. In this review, I will focus on the delicate and important balance between stability and plasticity at telomeres and subtelomeres of a kinetoplastid parasite, Trypanosoma brucei, which causes human African trypanosomiasis and undergoes antigenic variation to evade the host immune response. I will summarize the current understanding about T. brucei telomere protein complex, the telomeric transcript, and telomeric R-loops, focusing on their roles in maintaining telomere and subtelomere stability and integrity. The similarities and differences in functions and underlying mechanisms of T. brucei telomere factors will be compared with those in human and yeast cells.

ABSTRACT RAP1 is a telomere protein that is well conserved from protozoa to mammals. It plays important roles in chromosome end protection, telomere length control, and gene expression/silencing at both telomeric and nontelomeric loci. Interaction with different partners is an important mechanism by which RAP1 executes its different functions in yeast. The RAP1 ortholog in Trypanosoma brucei is essential for variant surface glycoprotein (VSG) monoallelic expression, an important aspect of antigenic variation, where T. brucei regularly switches its major surface antigen, VSG, to evade the host immune response. Like other RAP1 orthologs, T. brucei RAP1 (TbRAP1) has conserved functional domains, including BRCA1 C terminus (BRCT), Myb, MybLike, and RAP1 C terminus (RCT). To study functions of various TbRAP1 domains, we established a strain in which one endogenous allele of TbRAP1 is flanked by loxP repeats, enabling its conditional deletion by Cre-mediated recombination. We replaced the other TbRAP1 allele with various mutant alleles lacking individual functional domains and examined their nuclear localization and protein interaction abilities. The N terminus, BRCT, and RCT of TbRAP1 are required for normal protein levels, while the Myb and MybLike domains are essential for normal cell growth. Additionally, the Myb domain of TbRAP1 is required for its interaction with T. brucei TTAGGG repeat-binding factor (TbTRF), while the BRCT domain is required for its self-interaction. Furthermore, the TbRAP1 MybLike domain contains a bipartite nuclear localization signal that is required for its interaction with importin α and its nuclear localization. Interestingly, RAP1’s self-interaction and the interaction between RAP1 and TRF are conserved from kinetoplastids to mammals. However, details of the interaction interfaces have changed throughout evolution. IMPORTANCE Trypanosoma brucei causes human African trypanosomiasis and regularly switches its major surface antigen, VSG, to evade the host immune response. VSGs are expressed from subtelomeres in a monoallelic fashion. TbRAP1, a telomere protein, is essential for cell viability and VSG monoallelic expression and suppresses VSG switching. Although TbRAP1 has conserved functional domains in common with its orthologs from yeasts to mammals, the domain functions are unknown. RAP1 orthologs have pleiotropic functions, and interaction with different partners is an important means by which RAP1 executes its different roles. We have established a Cre-loxP-mediated conditional knockout system for TbRAP1 and examined the roles of various functional domains in protein expression, nuclear localization, and protein-protein interactions. This system enables further studies of TbRAP1 point mutation phenotypes. We have also determined functional domains of TbRAP1 that are required for several different protein interactions, shedding light on the underlying mechanisms of TbRAP1-mediated VSG silencing.

Trypanosoma brucei causes human African trypanosomiasis and regularly switches its major surface antigen, Variant Surface Glycoprotein (VSG), to evade the host immune response. Such antigenic variation is a key pathogenesis mechanism that enables T. brucei to establish long-term infections. VSG is expressed exclusively from subtelomere loci in a strictly monoallelic manner, and DNA recombination is an important VSG switching pathway. The integrity of telomere and subtelomere structure, maintained by multiple telomere proteins, is essential for T. brucei viability and for regulating the monoallelic VSG expression and VSG switching. Here we will focus on T. brucei TRF and RAP1, two telomere proteins with unique nucleic acid binding activities, and summarize their functions in telomere integrity and stability, VSG switching, and monoallelic VSG expression. Targeting the unique features of TbTRF and TbRAP1′s nucleic acid binding activities to perturb the integrity of telomere structure and disrupt VSG monoallelic expression may serve as potential therapeutic strategy against T. brucei.

Bacterial communities in gastrointestinal tracts (GIT) play an important role in animal health and performance. Despite its importance, little information is available on the establishment of microbial populations in the goat GIT or on changes occurring during early development. Therefore, this study investigated the bacterial community dynamics of the rumen, duodenum, jejunum, ileum, cecum, and colon in 15 goats at five developmental stages (0, 14, 28, 42, and 56 days old) by using 16S rDNA sequencing and quantitative real-time PCR technology. 940 genera were found to belong to 44 phyla distributed along the GIT. As a whole, the microbial richness and diversity showed a clear increasing trend as the kids aged and alpha diversity differed significantly among GIT compartments mainly occurring at middle day ages (14 and 28 days). Principal coordinate analysis indicated that the bacterial community displayed distinct temporal and spatial specificity along the GIT in preweaning goats. As kids aged, the phylum Firmicutes was replaced by Bacteroidetes in rumen, whereas Proteobacteria in the large intestine was displaced by Firmicutes. The phylum Proteobacteria was mainly present in the small intestine in older animals. In the rumen, taxa, such as Bacillus and Lactococcus decreased and Prevotella, Treponema, Ruminococcus, and unclassified Prevotellaceae increased with the age of kids. Furthermore, a lower proportion of taxa, such as Lactobacillus and Bacteroides was observed with higher abundances of both Christensenellaceae_R_7 and Ruminococcus in duodenum and jejunum in older animals. In the large intestine, the microbiota displayed taxonomic dynamics with increases of Ruminococcaceae UCG 005, unclassified Lachnospiraceae, Barnesiella, and Blautia as kids aged. Predicted pathway analysis suggested that genes involved in amino acid metabolism, and translation were abundant in both rumen and duodenum, while genes involved in membrane transport and carbohydrate metabolism were enriched in the large intestine. These results indicate that both the microbial colonization process and potential function exert a temporal-spatial specificity throughout the GIT of goats. This study provides new insight into the temporal dynamics of GIT microbiota development during preweaning and will aid to develop strategies for improving animal health and downstream production.

Ruminant animals possess a characteristic four-compartment stomach (rumen, reticulum, omasum, and abomasum) that is specialized for pre-intestinal digestion of plant materials. Of these four compartments, the rumen is the largest. The rumen’s diverse microbial community has been well studied. However, the current understanding of microbial profiles in the reticulum, omasum and abomasum are lacking. In the present study, fluid samples from the reticulum, omasum, and abomasum of goats at 3, 7, 14, 21, 28, 42, and 56 days after birth, as well as the negative controls (NC) used for microbial DNA extraction, were subjected to 16S rRNA sequencing. By filtering operational taxonomic units (OTUs) in NC, distinct temporal distributions of microbes were observed in the different compartments, we showed that the OTUs in control samples had a large effect to the samples with low microbial density. In addition, Proteobacteria gradually decreased with age from days 3 to 56 in all three compartments, and the relative abundance of Bacteroidetes increased from 24.15% (Day 3) to 52.03% (Day 56) in abomasum. Network analysis revealed that Prevotellaceae_UGG-03 and Rikenellaceae_RC9 were positively correlated with Prevotella_1, lending support to the well understood fact that cellulose is well digested in compound stomachs prior to the rumen. Pathway analysis revealed that gene expression in abomasum at Day 3 were primarily related to Glycolysis/Gluconeogenesis and Pyruvate metabolism, suggesting that colostrum digestion is the dominant function of the abomasum at an early age. These findings combined with other recent rumen microbiota data show that the microbiome landscape represents three distinct stages in ruminant stomachs. The first stage is to gain access to external microorganisms at Day 0–14, the secondary stage is for microbial transition at Day 14–28, and the third stage is for exogenous and endogenous microbial colonization beyond Day 28 of age. Our results provide insight into microbiota dynamics in ruminant stomachs, and will facilitate efforts for the maintenance of gastrointestinal balance and intervention with starter diets in juvenile ruminants during early development.

Trypanosoma brucei causes debilitating human African trypanosomiasis and evades the host's immune response by regularly switching its major surface antigen, VSG, which is expressed exclusively from subtelomeric loci. We previously showed that two interacting telomere proteins, TbTRF and TbTIF2, are essential for cell proliferation and suppress VSG switching by inhibiting DNA recombination events involving the whole active VSG expression site. We now find that TbTIF2 stabilizes TbTRF protein levels by inhibiting their degradation by the 26S proteasome, indicating that decreased TbTRF protein levels in TbTIF2-depleted cells contribute to more frequent VSG switching and eventual cell growth arrest. Surprisingly, although TbTIF2 depletion leads to more subtelomeric DNA double strand breaks (DSBs) that are both potent VSG switching inducers and detrimental to cell viability, TbTRF depletion does not increase the amount of DSBs inside subtelomeric VSG expression sites. Furthermore, expressing an ectopic allele of F2H-TbTRF in TbTIF2 RNAi cells allowed cells to maintain normal TbTRF protein levels for a longer frame of time. This resulted in a mildly better cell growth and partially suppressed the phenotype of increased VSG switching frequency but did not suppress the phenotype of more subtelomeric DSBs in TbTIF2-depleted cells. Therefore, TbTIF2 depletion has two parallel effects: decreased TbTRF protein levels and increased subtelomeric DSBs, both resulting in an acute increased VSG switching frequency and eventual cell growth arrest.

OBJECTIVES:There is an urgent need to develop a safe, effective, orally active, and inexpensive therapy for African trypanosomiasis due to the drawbacks of current drugs. Selective tubulin inhibitors have the potential to be promising drug candidates for the treatment of this disease, which is based on the tubulin protein structural difference between mammalian and trypanosome cells. We propose to identify novel tubulin inhibitors from a compound library developed based on the lead compounds that selectively target trypanosomiasis. METHODS:We used Trypanosoma brucei brucei as the parasite model, and human normal kidney cells and mouse microphage cells as the host model. Growth rates of both trypanosomes and mammalian cells were determined as a means to screen compounds that selectively inhibit the proliferation of parasites. Furthermore, we examined the cell cycle profile of the parasite and compared tubulin polymerization dynamics before and after the treatment using identified compounds. Last, in vivo anti-parasite activities of these compounds were determined in T. brucei-infected mice. RESULTS:Three compounds were selected that are 100 fold more effective against the growth of T. brucei cells than mammalian cells. These compounds caused cell cycle progression defects in T. brucei cells. Western analyses indicated that these compounds decreased tubulin polymerization in T. brucei cells. The in vivo investigation revealed that these compounds, when admitted orally, inhibited T. brucei cell proliferation in mouse blood. However, they were not potent enough to clear up the infection completely. CONCLUSIONS:These compounds are promising lead compounds as orally active agents for drug development of anti-trypanosome agents. A more detail structure activity relationship (SAR) was summarized that will be used to guide future lead optimization to improve the selectivity and potency of the current compounds.

The Hedgehog (HH) signaling pathway is critical for normal embryonic development, tissue patterning and cell differentiation. Aberrant HH signaling is involved in multiple human cancers. HH signaling involves a multi-protein cascade activating the GLI proteins that transcriptionally regulate HH target genes. We have previously reported that HH signaling is essential for human colon cancer cell survival and inhibition of this signal induces DNA damage and extensive cell death. Here we report that the HH/GLI axis regulates human telomerase reverse transcriptase (hTERT), which determines the replication potential of cancer cells. Suppression of GLI1/GLI2 functions by a C-terminus truncated GLI3 repressor mutant (GLI3R), or by GANT61, a pharmacological inhibitor of GLI1/GLI2, reduced hTERT protein expression in human colon cancer, prostate cancer and Glioblastoma multiforme (GBM) cell lines. Expression of an N-terminus deleted constitutively active mutant of GLI2 (GLI2ΔN) increased hTERT mRNA and protein expression and hTERT promoter driven luciferase activity in human colon cancer cells while GANT61 inhibited hTERT mRNA expression and hTERT promoter driven luciferase activity. Chromatin immunoprecipitation with GLI1 or GLI2 antibodies precipitated fragments of the hTERT promoter in human colon cancer cells, which was reduced upon exposure to GANT61. In contrast, expression of GLI1 or GLI2ΔN in non-malignant 293T cells failed to alter the levels of hTERT mRNA and protein, or hTERT promoter driven luciferase activity. Further, expression of GLI2ΔN increased the telomerase enzyme activity, which was reduced by GANT61 administration in human colon cancer, prostate cancer, and GBM cells. These results identify hTERT as a direct target of the HH signaling pathway, and reveal a previously unknown role of the HH/GLI axis in regulating the replication potential of cancer cells. These findings are of significance in understanding the important regulatory mechanisms that determine the functions of HH/GLI signaling in cancer cells.

miR-219-5p has been reported to act as either a tumor suppressor or a tumor promoter in different cancers by targeting different genes. In the present study, we demonstrated that miR-219-5p negatively regulated the expression of TBXT, a known epithelial-mesenchymal transition (EMT) inducer, by directly binding to TBXT 3'-untranslated region. As a result of its inhibition on TBXT expression, miR-219-5p suppressed EMT and cell migration and invasion in breast cancer cells. The re-introduction of TBXT in miR-219-5p overexpressing cells decreased the inhibitory effects of miR-219 on EMT and cell migration and invasion. Moreover, miR-219-5p decreased breast cancer stem cell (CSC) marker genes expression and reduced the mammosphere forming capability of cells. Overall, our study highlighted that TBXT is a novel target of miR-219-5p. By suppressing TBXT, miR-219-5p plays an important role in EMT and cell migration and invasion of breast cancer cells. [ABSTRACT FROM AUTHOR]