Trypanosoma brucei ribonuclease H2A is an essential R-loop processing enzyme whose loss causes DNA damage during transcription initiation and antigenic variation.

Ribonucleotides represent a threat to DNA genome stability and transmission. Two types of Ribonuclease H (RNase H) excise ribonucleotides when they form part of the DNA strand, or hydrolyse RNA when it base-pairs with DNA in structures termed R-loops. Loss of either RNase H is lethal in mammals, whereas yeast survives the absence of both enzymes. RNase H1 loss is tolerated by the parasite Trypanosoma brucei but no work has examined the function of RNase H2. Here we show that loss of T. brucei RNase H2 (TbRH2A) leads to growth and cell cycle arrest that is concomitant with accumulation of nuclear damage at sites of RNA polymerase (Pol) II transcription initiation, revealing a novel and critical role for RNase H2. Differential gene expression analysis reveals limited overall changes in RNA levels for RNA Pol II genes after TbRH2A loss, but increased perturbation of nucleotide metabolic genes. Finally, we show that TbRH2A loss causes R-loop and DNA damage accumulation in telomeric RNA Pol I transcription sites, also leading to altered gene expression. Thus, we demonstrate separation of function between two nuclear T. brucei RNase H enzymes during RNA Pol II transcription, but overlap in function during RNA Pol I-mediated gene expression during host immune evasion.

Genome-wide mapping reveals conserved and diverged R-loop activities in the unusual genetic landscape of the African trypanosome genome.

R-loops are stable RNA-DNA hybrids that have been implicated in transcription initiation and termination, as well as in telomere maintenance, chromatin formation, and genome replication and instability. RNA Polymerase (Pol) II transcription in the protozoan parasite Trypanosoma brucei is highly unusual: virtually all genes are co-transcribed from multigene transcription units, with mRNAs generated by linked trans-splicing and polyadenylation, and transcription initiation sites display no conserved promoter motifs. Here, we describe the genome-wide distribution of R-loops in wild type mammal-infective T. brucei and in mutants lacking RNase H1, revealing both conserved and diverged functions. Conserved localization was found at centromeres, rRNA genes and retrotransposon-associated genes. RNA Pol II transcription initiation sites also displayed R-loops, suggesting a broadly conserved role despite the lack of promoter conservation or transcription initiation regulation. However, the most abundant sites of R-loop enrichment were within the regions between coding sequences of the multigene transcription units, where the hybrids coincide with sites of polyadenylation and nucleosome-depletion. Thus, instead of functioning in transcription termination the most widespread localization of R-loops in T. brucei suggests a novel correlation with pre-mRNA processing. Finally, we find little evidence for correlation between R-loop localization and mapped sites of DNA replication initiation.

Genome-wide and protein kinase-focused RNAi screens reveal conserved and novel damage response pathways in Trypanosoma brucei.

All cells are subject to structural damage that must be addressed for continued growth. A wide range of damage affects the genome, meaning multiple pathways have evolved to repair or bypass the resulting DNA lesions. Though many repair pathways are conserved, their presence or function can reflect the life style of individual organisms. To identify genome maintenance pathways in a divergent eukaryote and important parasite, Trypanosoma brucei, we performed RNAi screens to identify genes important for survival following exposure to the alkylating agent methyl methanesulphonate. Amongst a cohort of broadly conserved and, therefore, early evolved repair pathways, we reveal multiple activities not so far examined functionally in T. brucei, including DNA polymerases, DNA helicases and chromatin factors. In addition, the screens reveal Trypanosoma- or kinetoplastid-specific repair-associated activities. We also provide focused analyses of repair-associated protein kinases and show that loss of at least nine, and potentially as many as 30 protein kinases, including a nuclear aurora kinase, sensitises T. brucei to alkylation damage. Our results demonstrate the potential for synthetic lethal genome-wide screening of gene function in T. brucei and provide an evolutionary perspective on the repair pathways that underpin effective responses to damage, with particular relevance for related kinetoplastid pathogens. By revealing that a large number of diverse T. brucei protein kinases act in the response to damage, we expand the range of eukaryotic signalling factors implicated in genome maintenance activities.

Vacuolar ATPase depletion affects mitochondrial ATPase function, kinetoplast dependency, and drug sensitivity in trypanosomes.

Kinetoplastid parasites cause lethal diseases in humans and animals. The kinetoplast itself contains the mitochondrial genome, comprising a huge, complex DNA network that is also an important drug target. Isometamidium, for example, is a key veterinary drug that accumulates in the kinetoplast in African trypanosomes. Kinetoplast independence and isometamidium resistance are observed where certain mutations in the F1-γ-subunit of the two-sector F1Fo-ATP synthase allow for Fo-independent generation of a mitochondrial membrane potential. To further explore kinetoplast biology and drug resistance, we screened a genome-scale RNA interference library in African trypanosomes for isometamidium resistance mechanisms. Our screen identified 14 V-ATPase subunits and all 4 adaptin-3 subunits, implicating acidic compartment defects in resistance; V-ATPase acidifies lysosomes and related organelles, whereas adaptin-3 is responsible for trafficking among these organelles. Independent strains with depleted V-ATPase or adaptin-3 subunits were isometamidium resistant, and chemical inhibition of the V-ATPase phenocopied this effect. While drug accumulation in the kinetoplast continued after V-ATPase subunit depletion, acriflavine-induced kinetoplast loss was specifically tolerated in these cells and in cells depleted for adaptin-3 or endoplasmic reticulum membrane complex subunits, also identified in our screen. Consistent with kinetoplast dispensability, V-ATPase defective cells were oligomycin resistant, suggesting ATP synthase uncoupling and bypass of the normal Fo-A6-subunit requirement; this subunit is the only kinetoplast-encoded product ultimately required for viability in bloodstream-form trypanosomes. Thus, we describe 30 genes and 3 protein complexes associated with kinetoplast-dependent growth. Mutations affecting these genes could explain natural cases of dyskinetoplasty and multidrug resistance. Our results also reveal potentially conserved communication between the compartmentalized two-sector rotary ATPases.

Minimally Invasive Plate Osteosynthesis for Treatment of Ankle Fractures in High-Risk Patients.

Wound healing problems are the most common complication after open reduction with internal fixation (ORIF) of unstable ankle fractures. The incidence is especially high among elderly patients with medical comorbidities and patients with compromised soft tissues. Minimally invasive plate osteosynthesis (MIPO) might provide a safer alternative to ORIF by preventing extensive soft tissue dissection and preserving the blood supply. We conducted a retrospective review of 44 consecutive patients who had undergone MIPO of unstable ankle fractures. All patients had a minimum 1-year follow-up (mean 82 weeks); 80% were aged ≥60 years, 52% had diabetes, and 45% had a compromised soft tissue envelope. Immediate postoperative radiographs were evaluated for the quality of reduction, and clinical records were analyzed for the complication rate. Good to excellent anatomic reduction was achieved in 89% of the patients. The overall complication rate was 27%, including 25% surgical wound dehiscence, 9% infection, and 11% loss of reduction. No patient experienced nerve injury. Those with a history of ankle fracture dislocation and a compromised soft tissue envelope preoperatively had a significantly greater incidence of surgical wound dehiscence and complications overall compared with those without (p = .016 and p = .035; p = .045 and p = .009, respectively). Peripheral vascular disease was a statistically significant predictor of surgical wound dehiscence (p = .010). The overall complication rate in our study was comparable to that seen in similar populations treated with conventional ORIF. In conclusion, our results suggest that MIPO in high-risk patients is a safe alternative, with predictable outcomes, comparable to those of traditional open techniques.

Smooth Muscle Enriched Long Noncoding RNA (SMILR) Regulates Cell Proliferation.

BACKGROUND: Phenotypic switching of vascular smooth muscle cells from a contractile to a synthetic state is implicated in diverse vascular pathologies, including atherogenesis, plaque stabilization, and neointimal hyperplasia. However, very little is known about the role of long noncoding RNA (lncRNA) during this process. Here, we investigated a role for lncRNAs in vascular smooth muscle cell biology and pathology. METHODS AND RESULTS: Using RNA sequencing, we identified >300 lncRNAs whose expression was altered in human saphenous vein vascular smooth muscle cells following stimulation with interleukin-1α and platelet-derived growth factor. We focused on a novel lncRNA (Ensembl: RP11-94A24.1), which we termed smooth muscle-induced lncRNA enhances replication (SMILR). Following stimulation, SMILR expression was increased in both the nucleus and cytoplasm, and was detected in conditioned media. Furthermore, knockdown of SMILR markedly reduced cell proliferation. Mechanistically, we noted that expression of genes proximal to SMILR was also altered by interleukin-1α/platelet-derived growth factor treatment, and HAS2 expression was reduced by SMILR knockdown. In human samples, we observed increased expression of SMILR in unstable atherosclerotic plaques and detected increased levels in plasma from patients with high plasma C-reactive protein. CONCLUSIONS: These results identify SMILR as a driver of vascular smooth muscle cell proliferation and suggest that modulation of SMILR may be a novel therapeutic strategy to reduce vascular pathologies.

Cardiac Hypertrophy Is Inhibited by a Local Pool of cAMP Regulated by Phosphodiesterase 2.

RATIONALE: Chronic elevation of 3'-5'-cyclic adenosine monophosphate (cAMP) levels has been associated with cardiac remodeling and cardiac hypertrophy. However, enhancement of particular aspects of cAMP/protein kinase A signaling seems to be beneficial for the failing heart. cAMP is a pleiotropic second messenger with the ability to generate multiple functional outcomes in response to different extracellular stimuli with strict fidelity, a feature that relies on the spatial segregation of the cAMP pathway components in signaling microdomains. OBJECTIVE: How individual cAMP microdomains affect cardiac pathophysiology remains largely to be established. The cAMP-degrading enzymes phosphodiesterases (PDEs) play a key role in shaping local changes in cAMP. Here we investigated the effect of specific inhibition of selected PDEs on cardiac myocyte hypertrophic growth. METHODS AND RESULTS: Using pharmacological and genetic manipulation of PDE activity, we found that the rise in cAMP resulting from inhibition of PDE3 and PDE4 induces hypertrophy, whereas increasing cAMP levels via PDE2 inhibition is antihypertrophic. By real-time imaging of cAMP levels in intact myocytes and selective displacement of protein kinase A isoforms, we demonstrate that the antihypertrophic effect of PDE2 inhibition involves the generation of a local pool of cAMP and activation of a protein kinase A type II subset, leading to phosphorylation of the nuclear factor of activated T cells. CONCLUSIONS: Different cAMP pools have opposing effects on cardiac myocyte cell size. PDE2 emerges as a novel key regulator of cardiac hypertrophy in vitro and in vivo, and its inhibition may have therapeutic applications.

Diagnostic and Therapeutic Applications of Quantum Dots in Nanomedicine.

The interest in Quantum Dots as a class of nanomaterials has grown considerably since their discovery by Ekimov and Efros in the early 1980s. Although this early work focussed primarily on CdSe-based nanocrystals, the field has now expanded to include various classes of nanoparticles with different types of core, shell or passivation chemistry. Such differences can have a profound effect on the optical properties and potential biocompatibility of the resulting constructs. Although QDs have predominantly been used for imaging and sensing applications, more examples of their use as therapeutics are beginning to emerge. In this chapter we discuss the progress made over the past decade in developing QDs for imaging and therapeutic applications.

Optical probes for the detection of protons, and alkali and alkaline earth metal cations.

Luminescent sensors and switches continue to play a key role in shaping our understanding of key biochemical processes, assist in the diagnosis of disease and contribute to the design of new drugs and therapies. Similarly, their contribution to the environment cannot be understated as they offer a portable means to undertake field testing for hazardous chemicals and pollutants such as heavy metals. From a physiological perspective, the Group I and II metal ions are among the most important in the periodic table with blood plasma levels of H(+), Na(+) and Ca(2+) being indicators of several possible disease states. In this review, we examine the progress that has been made in the development of luminescent probes for Group I and Group II ions as well as protons. The potential applications of these probes and the mechanism involved in controlling their luminescent response upon analyte binding will also be discussed.

Effects of the lapidus arthrodesis and chevron bunionectomy on plantar forefoot pressures.

Hallux valgus with or without first ray insufficiency has been strongly implicated as a contributing factor in lesser metatarsal overload. The principle goals of a bunionectomy are to relieve the pain, correct the deformity, and restore first metatarsophalangeal joint congruity. Until now, little evidence has been available to assess the effects of bunionectomy procedures on forefoot pressure. The primary aim of the present prospective study was to evaluate the preoperative and postoperative plantar pressures after 2 specific bunionectomies: the chevron bunionectomy and Lapidus arthrodesis. A total of 68 subjects, 34 in each group, were included for radiographic and pedographic evaluation. Both procedures demonstrated radiographic improvements in the mean intermetatarsal and hallux abductus angles. The mean hallux plantar pressure decreased significantly in both procedure groups (p < .001). However, Lapidus group exhibited an increase in the mean fifth metatarsal head plantar pressure (p = .008) and pressure under the fifth metatarsal as a percentage of the total forefoot pressure (p = .01). Furthermore, the pressure under the second metatarsal as a percentage of the total forefoot pressure decreased significantly (p = .01). This study suggests that the Lapidus arthrodesis and chevron bunionectomy both provide correction for hallux valgus deformity, but when comparing forefoot load sharing pressures, the Lapidus arthrodesis appeared to have greater influence on the load sharing distribution of forefoot pressure than did the bunionectomy employing the chevron osteotomy.

Management of Talus Fractures.

Fractures of the talus are life-changing events. The talus is of vital importance to normal gait. Given its importance, great care is needed in diagnosing and treating these injuries. The threshold for operative treatment and accurate anatomic reduction should be low. Surgical tenets include the avoidance of extensive subperiosteal dissection to minimize vascular disruption. The complications with injuries to the talus are extensive and include avascular necrosis (AVN). Although AVN can prove to be a devastating sequela from this injury, it occurs less frequently than posttraumatic arthritis.

SEC31A may be associated with pituitary hormone deficiency and gonadal dysgenesis.

PURPOSE: Disorders/differences of sex development (DSD) result from variants in many different human genes but, frequently, have no detectable molecular cause. METHODS: Detailed clinical and genetic phenotyping was conducted on a family with three children. A Sec31a animal model and functional studies were used to investigate the significance of the findings. RESULTS: By trio whole-exome DNA sequencing we detected a heterozygous de novo nonsense SEC31A variant, in three children of healthy non-consanguineous parents. The children had different combinations of disorders that included complete gonadal dysgenesis and multiple pituitary hormone deficiency. SEC31A encodes a component of the COPII coat protein complex, necessary for intracellular anterograde vesicle-mediated transport between the endoplasmic reticulum (ER) and Golgi. CRISPR-Cas9 targeted knockout of the orthologous Sec31a gene region resulted in early embryonic lethality in homozygous mice. mRNA expression of ER-stress genes ATF4 and CHOP was increased in the children, suggesting defective protein transport. The pLI score of the gene, from gnomAD data, is 0.02. CONCLUSIONS: SEC31A might underlie a previously unrecognised clinical syndrome comprising gonadal dysgenesis, multiple pituitary hormone deficiencies, dysmorphic features and developmental delay. However, a variant that remains undetected, in a different gene, may alternatively be causal in this family.

cGMP signals modulate cAMP levels in a compartment-specific manner to regulate catecholamine-dependent signaling in cardiac myocytes.

RATIONALE: cAMP and cGMP are intracellular second messengers involved in heart pathophysiology. cGMP can potentially affect cAMP signals via cGMP-regulated phosphodiesterases (PDEs). OBJECTIVE: To study the effect of cGMP signals on the local cAMP response to catecholamines in specific subcellular compartments. METHODS AND RESULTS: We used real-time FRET imaging of living rat ventriculocytes expressing targeted cAMP and cGMP biosensors to detect cyclic nucleotides levels in specific locales. We found that the compartmentalized, but not the global, cAMP response to isoproterenol is profoundly affected by cGMP signals. The effect of cGMP is to increase cAMP levels in the compartment where the protein kinase (PK)A-RI isoforms reside but to decrease cAMP in the compartment where the PKA-RII isoforms reside. These opposing effects are determined by the cGMP-regulated PDEs, namely PDE2 and PDE3, with the local activity of these PDEs being critically important. The cGMP-mediated modulation of cAMP also affects the phosphorylation of PKA targets and myocyte contractility. CONCLUSIONS: cGMP signals exert opposing effects on local cAMP levels via different PDEs the activity of which is exerted in spatially distinct subcellular domains. Inhibition of PDE2 selectively abolishes the negative effects of cGMP on cAMP and may have therapeutic potential.

Complications associated with foot and ankle arthroscopy.

Despite a late start within the realm of arthroscopy, foot and ankle arthroscopy proves to be an important diagnostic and treatment tool for the foot and ankle specialist. As indication for arthroscopy increases, complications associated with foot and ankle arthroscopy must be revisited. We reviewed 405 foot and ankle arthroscopic procedures performed on 390 patients in 4 different facilities over a 3-year period extending from January 2005 to August 2008. Two-hundred-sixty foot and ankle arthroscopic procedures on 251 patients met our inclusion criteria. A total of 246 ankle and 14 posterior subtalar arthroscopic procedures were performed with a mean follow-up of 10.7 ± 3.5 months. Patient demographics, preoperative findings, intraoperative technique, and postoperative course were reviewed. We failed to identify statistically significant predictive factors for complications. Arthroscopy performed in combination with adjunctive procedures showed a trend toward higher complication rate, although statistical significance was not noted. Overall, 20 cases (7.69%) experienced arthroscopy-related complications, and this finding was comparable with previously published results. The most common complication was cutaneous nerve injury, which involved 9 cases (3.46%), and localized superficial infection, which involved 8 cases (3.08%). Injury to the superficial peroneal nerve accounted for 5 of the cutaneous nerve injuries. There were no cases of arthroscopy-related vascular injury. All cases of superficial postoperative infection resolved with antibiotic therapy, and none of the cases required return to the operating room. These results were also similar to published data.

Association between ankle fractures and obesity.

Obesity is an epidemic in the United States and is associated with an increased risk of musculoskeletal problems. Rotational injuries of the ankle with a Weber C fibula fracture have a greater risk of syndesmosis disruption and instability. The goal of the present study was to explore the association between obesity and ankle fractures. Using a retrospective review, the radiographs of 280 patients with an ankle fracture were reviewed and classified using the Weber classification, which was then associated with the body mass index, gender, age, diabetes, tobacco use, and osteoporosis. Patients with a body mass index of 30 kg/m(2) or greater (odds ratio 1.78), men (odds ratio 1.74), and age 25 years or younger (odds ratio 3.97) had greater odds of having a Weber C ankle fracture (compared with Weber A and B) and Weber C and B (compared with Weber A). Diabetes mellitus, osteoporosis/osteopenia, and current tobacco use were not significantly associated with the severity of the ankle fracture. The results from the present study suggest that obesity presents a greater risk of sustaining a more proximal distal fibula fracture.

Complications after open reduction and internal fixation of ankle fractures in the elderly.

BACKGROUND: Open reduction with internal fixation for unstable ankle fractures is relatively predictable with excellent outcomes. However, the management of ankle fractures in the elderly remains less predictable secondary to the various co-morbidities associated with advanced age. METHODS: A retrospective chart review of 216 patients over the age of 60 that sustained an ankle fracture, was performed to determine the incidence of complications after ORIF of ankle fractures in an elderly population in the perioperative course. Secondly, the incidence of complications in patients that had locking plate fixation compared to those that had non-locking plate fixation was determined. Lastly, the effect of early weight bearing on the incidence of complications was analyzed. RESULTS: There was not a statistically significant difference in the complication rates between the group with co-morbidities (19.01%) and those without (11.96%). The postoperative complication with the highest incidence was wound dehiscence (9.7%), and only diabetes significantly predicted wound dehiscence. The fixation construct and weight-bearing protocol failed to significantly predict any of the indexed complications. CONCLUSIONS: Overall, the results suggest that surgical treatment of unstable ankle fractures in the elderly is fairly predictable with an acceptable complication rate. The complication rates are higher with increased age and diabetes, but they failed to reach statistical significance. Conventional plating appears to provide adequate stability without increased risk of hardware failure. In addition patients that were allowed to walk within the first 2 weeks postoperatively did not experience a higher rate of hardware failure.

Transcriptome analyses of nine endocrine tissues identifies organism-wide transcript distribution and structure in the Siberian hamster.

Temperate zone animals exhibit seasonal variation in multiple endocrine systems. In most cases, peripheral organs display robust switches in tissue involution and recrudescence in mass. Our understanding of the molecular control of tissue-specific changes in seasonal function remains limited. Central to this problem is the lack of information on the nucleic acid structure, and distribution of transcripts across tissues in seasonal model organisms. Here we report the transcriptome profile of nine endocrine tissues from Siberian hamsters. Luteinizing hormone receptor expression was localized to gonadal tissues and confirmed previous distribution analyses. Assessment of the prolactin receptor reveal relatively high abundance across tissues involved in reproduction, energy, and water homeostasis. Neither melatonin receptor-1a, nor -1b, were found to be expressed in most tissues. Instead, the closely related G-protein coupled receptor Gpr50 was widely expressed in peripheral tissues. Epigenetic enzymes such as DNA methyltransferase 3a, was widely expressed and the predominant DNA methylation enzyme. Quantitative PCR analyses revealed some sex- and tissue-specific differences for prolactin receptor and DNA methyltransferase 3a expression. These data provide significant information on the distribution of transcripts, relative expression levels and nucleic acid sequences that will facilitate molecular studies into the seasonal programs in mammalian physiology.

Radiographic outcomes of adult acquired flatfoot corrected by medial column arthrodesis with or without a medializing calcaneal osteotomy.

Medial column arthrodesis and calcaneal osteotomies are commonly used for adult acquired flatfoot surgical reconstruction. In this retrospective study, 41 patients (47 feet) with a mean age of 55 ± 13.5 years underwent a medial column arthrodesis, with or without calcaneal osteotomy, between 1999 and 2007. The indication for surgery was a painful flatfoot deformity with peritalar subluxation, and a fault in the naviculocuneiform joint. At a mean of 9.6 (range 3-43) months postoperatively, in patients who underwent a medial column arthrodesis, radiographs showed a mean decrease in the talonavicular coverage angle of 10.2° ± 8.7° (P < .001), and mean increases in the lateral talometatarsal and calcaneal inclination angle of 10.7° ± 5.1° (P < .001) and of 3.2° ± 2.7° (P < .001), respectively. In patients who underwent a combined medial column arthrodesis and a medializing calcaneal osteotomy, the talonavicular coverage angle decreased by a mean of 12.1° ± 6.1° (P < .001), while the lateral talometatarsal angle and calcaneal inclination angle increased by a mean of 12.3° ± 6.1° (P < .001) and 3.1° ± 2.7° (P < .001), respectively, from preoperative values. Four nonunions (4 of 47, 8.51%) occurred at the naviculocuneiform joint and 1 nonunion (1 of 32, 3.13%) occurred at the tarsometatarsal joint. These findings demonstrate marked improvement of radiographic flatfoot parameters following a medial column arthrodesis with or without a medializing calcaneal osteotomy.

Gene Regulation Network Analysis on Human Prostate Orthografts Highlights a Potential Role for the JMJD6 Regulon in Clinical Prostate Cancer.

BACKGROUND: Prostate cancer (PCa) is the second most common tumour diagnosed in men. Tumoral heterogeneity in PCa creates a significant challenge to develop robust prognostic markers and novel targets for therapy. An analysis of gene regulatory networks (GRNs) in PCa may provide insight into progressive PCa. Herein, we exploited a graph-based enrichment score to integrate data from GRNs identified in preclinical prostate orthografts and differentially expressed genes in clinical resected PCa. We identified active regulons (transcriptional regulators and their targeted genes) associated with PCa recurrence following radical prostatectomy. METHODS: The expression of known transcription factors and co-factors was analysed in a panel of prostate orthografts (n = 18). We searched for genes (as part of individual GRNs) predicted to be regulated by the highest number of transcriptional factors. Using differentially expressed gene analysis (on a per sample basis) coupled with gene graph enrichment analysis, we identified candidate genes and associated GRNs in PCa within the UTA cohort, with the most enriched regulon being JMJD6, which was further validated in two additional cohorts, namely EMC and ICGC cohorts. Cox regression analysis was performed to evaluate the association of the JMJD6 regulon activity with disease-free survival time in the three clinical cohorts as well as compared to three published prognostic gene signatures (TMCC11, BROMO-10 and HYPOXIA-28). RESULTS: 1308 regulons were correlated to transcriptomic data from the three clinical prostatectomy cohorts. The JMJD6 regulon was identified as the top enriched regulon in the UTA cohort and again validated in the EMC cohort as the top-ranking regulon. In both UTA and EMC cohorts, the JMJD6 regulon was significantly associated with cancer recurrence. Active JMJD6 regulon also correlated with disease recurrence in the ICGC cohort. Furthermore, Kaplan-Meier analysis confirmed shorter time to recurrence in patients with active JMJD6 regulon for all three clinical cohorts (UTA, EMC and ICGC), which was not the case for three published prognostic gene signatures (TMCC11, BROMO-10 and HYPOXIA-28). In multivariate analysis, the JMJD6 regulon status significantly predicted disease recurrence in the UTA and EMC, but not ICGC datasets, while none of the three published signatures significantly prognosticate for cancer recurrence. CONCLUSIONS: We have characterised gene regulatory networks from preclinical prostate orthografts and applied transcriptomic data from three clinical cohorts to evaluate the prognostic potential of the JMJD6 regulon.