Antidepressant drugs act by directly binding to TRKB neurotrophin receptors.

It is unclear how binding of antidepressant drugs to their targets gives rise to the clinical antidepressant effect. We discovered that the transmembrane domain of tyrosine kinase receptor 2 (TRKB), the brain-derived neurotrophic factor (BDNF) receptor that promotes neuronal plasticity and antidepressant responses, has a cholesterol-sensing function that mediates synaptic effects of cholesterol. We then found that both typical and fast-acting antidepressants directly bind to TRKB, thereby facilitating synaptic localization of TRKB and its activation by BDNF. Extensive computational approaches including atomistic molecular dynamics simulations revealed a binding site at the transmembrane region of TRKB dimers. Mutation of the TRKB antidepressant-binding motif impaired cellular, behavioral, and plasticity-promoting responses to antidepressants in vitro and in vivo. We suggest that binding to TRKB and allosteric facilitation of BDNF signaling is the common mechanism for antidepressant action, which may explain why typical antidepressants act slowly and how molecular effects of antidepressants are translated into clinical mood recovery.

DMC1 and RAD51 bind FxxA and FxPP motifs of BRCA2 via two separate interfaces.

In vertebrates, the BRCA2 protein is essential for meiotic and somatic homologous recombination due to its interaction with the RAD51 and DMC1 recombinases through FxxA and FxPP motifs (here named A- and P-motifs, respectively). The A-motifs present in the eight BRC repeats of BRCA2 compete with the A-motif of RAD51, which is responsible for its self-oligomerization. BRCs thus disrupt RAD51 nucleoprotein filaments in vitro. The role of the P-motifs is less studied. We recently found that deletion of Brca2 exons 12-14 encoding one of them (the prototypical 'PhePP' motif), disrupts DMC1 but not RAD51 function in mouse meiosis. Here we provide a mechanistic explanation for this phenotype by solving the crystal structure of the complex between a BRCA2 fragment containing the PhePP motif and DMC1. Our structure reveals that, despite sharing a conserved phenylalanine, the A- and P-motifs bind to distinct sites on the ATPase domain of the recombinases. The P-motif interacts with a site that is accessible in DMC1 octamers and nucleoprotein filaments. Moreover, we show that this interaction also involves the adjacent protomer and thus increases the stability of the DMC1 nucleoprotein filaments. We extend our analysis to other P-motifs from RAD51AP1 and FIGNL1.

GluK1 kainate receptors are necessary for functional maturation of parvalbumin interneurons regulating amygdala circuit function.

Parvalbumin expressing interneurons (PV INs) are key players in the local inhibitory circuits and their developmental maturation coincides with the onset of adult-type network dynamics in the brain. Glutamatergic signaling regulates emergence of the unique PV IN phenotype, yet the receptor mechanisms involved are not fully understood. Here we show that GluK1 subunit containing kainate receptors (KARs) are necessary for development and maintenance of the neurochemical and functional properties of PV INs in the lateral and basal amygdala (BLA). Ablation of GluK1 expression specifically from PV INs resulted in low parvalbumin expression and loss of characteristic high firing rate throughout development. In addition, we observed reduced spontaneous excitatory synaptic activity at adult GluK1 lacking PV INs. Intriguingly, inactivation of GluK1 expression in adult PV INs was sufficient to abolish their high firing rate and to reduce PV expression levels, suggesting a role for GluK1 in dynamic regulation of PV IN maturation state. The PV IN dysfunction in the absence of GluK1 perturbed the balance between evoked excitatory vs. inhibitory synaptic inputs and long-term potentiation (LTP) in LA principal neurons, and resulted in aberrant development of the resting-state functional connectivity between mPFC and BLA. Behaviorally, the absence of GluK1 from PV INs associated with hyperactivity and increased fear of novelty. These results indicate a critical role for GluK1 KARs in regulation of PV IN function across development and suggest GluK1 as a potential therapeutic target for pathologies involving PV IN malfunction.

Endothelial Responses to Curvature-Induced Flow Patterns in Engineered Cerebral Aneurysms.

Hemodynamic factors have long been associated with clinical outcomes in the treatment of cerebral aneurysms. Computational studies of cerebral aneurysm hemodynamics have provided valuable estimates of the mechanical environment experienced by the endothelium in both the parent vessel and aneurysmal dome walls and have correlated them with disease state. These computational-clinical studies have recently been correlated with the response of endothelial cells (EC) using either idealized or patient-specific models. Here, we present a robust workflow for generating anatomic-scale aneurysm models, establishing luminal cultures of ECs at physiological relevant flow profiles, and comparing EC responses to curvature mediated flow. We show that flow patterns induced by parent vessel curvature produce changes in wall shear stress (WSS) and wall shear stress gradients (WSSG) that are correlated with differences in cell morphology and cellular protein localization. Cells in higher WSS regions align better with the flow and display strong Notch1-extracellular domain (ECD) polarization, while, under low WSS, differences in WSSG due to curvature change were associated with less alignment and attenuation of Notch1-ECD polarization in ECs of the corresponding regions. These proof-of-concept results highlight the use of engineered cellularized aneurysm models for connecting computational fluid dynamics to the underlying endothelial biology that mediates disease.

Suicide as globalisation's Black Swan: global evidence.

OBJECTIVES: This empirical study investigated the relationship between globalisation and suicide rates. We examined whether there is a beneficial or harmful relationship between economic, political and social globalisation and the suicide rate. We also estimated whether this relationship differs in high-, middle- and low-income countries. STUDY DESIGN: Using panel data from 190 countries over the period 1990-2019, we examined the relationship between globalisation and suicide. METHOD: We compared the estimated effect of globalisation on suicide rates using robust fixed-effects models. Our results were robust to dynamic models and models with country-specific time trends. RESULTS: The effect of the KOF Globalisation Index on suicide was initially positive, leading to an increase in the suicide rate before decreasing. Concerning the effects of economic, political, and social dimensions of globalisation, we found a similar inverted U-shaped relationship. Unlike the middle-income and high-income countries, we found a U-shaped relationship for the case of low-income countries, indicating that suicide decreased with globalisation and then increased as globalisation continues to increase. Moreover, the effect of political globalisation disappeared in low-income countries. CONCLUSION: Policy-makers in high- and middle-income countries, below the turning points, and low-income countries, above the turning points, must protect vulnerable groups from globalisation's disruptive forces, which can increase social inequality. Consideration of local and global factors of suicide will potentially stimulate the development of measures that might reduce the suicide rate.

Pharmacological and optical activation of TrkB in Parvalbumin interneurons regulate intrinsic states to orchestrate cortical plasticity.

Elevated states of brain plasticity typical for critical periods of early postnatal life can be reinstated in the adult brain through interventions, such as antidepressant treatment and environmental enrichment, and induced plasticity may be critical for the antidepressant action. Parvalbumin-positive (PV) interneurons regulate the closure of developmental critical periods and can alternate between high and low plasticity states in response to experience in adulthood. We now show that PV plasticity states and cortical networks are regulated through the activation of TrkB neurotrophin receptors. Visual cortical plasticity induced by fluoxetine, a widely prescribed selective serotonin reuptake inhibitor (SSRI) antidepressant, was lost in mice with reduced expression of TrkB in PV interneurons. Conversely, optogenetic gain-of-function studies revealed that activation of an optically activatable TrkB (optoTrkB) specifically in PV interneurons switches adult cortical networks into a state of elevated plasticity within minutes by decreasing the intrinsic excitability of PV interneurons, recapitulating the effects of fluoxetine. TrkB activation shifted cortical networks towards a low PV configuration, promoting oscillatory synchrony, increased excitatory-inhibitory balance, and ocular dominance plasticity. OptoTrkB activation promotes the phosphorylation of Kv3.1 channels and reduces the expression of Kv3.2 mRNA providing a mechanism for the lower excitability. In addition, decreased expression and puncta of Synaptotagmin2 (Syt2), a presynaptic marker of PV interneurons involved in Ca2+-dependent neurotransmitter release, suggests lower inputs onto pyramidal neurons suppressing feed-forward inhibition. Together, the results provide mechanistic insights into how TrkB activation in PV interneurons orchestrates the activity of cortical networks and mediating antidepressant responses in the adult brain.

HSF2BP negatively regulates homologous recombination in DNA interstrand crosslink repair.

The tumor suppressor BRCA2 is essential for homologous recombination (HR), replication fork stability and DNA interstrand crosslink (ICL) repair in vertebrates. We show that ectopic production of HSF2BP, a BRCA2-interacting protein required for meiotic HR during mouse spermatogenesis, in non-germline human cells acutely sensitize them to ICL-inducing agents (mitomycin C and cisplatin) and PARP inhibitors, resulting in a phenotype characteristic of cells from Fanconi anemia (FA) patients. We biochemically recapitulate the suppression of ICL repair and establish that excess HSF2BP compromises HR by triggering the removal of BRCA2 from the ICL site and thereby preventing the loading of RAD51. This establishes ectopic expression of a wild-type meiotic protein in the absence of any other protein-coding mutations as a new mechanism that can lead to an FA-like cellular phenotype. Naturally occurring elevated production of HSF2BP in tumors may be a source of cancer-promoting genomic instability and also a targetable vulnerability.

Rapid herpes zoster infection on latissimus dorsi flap following breast reconstruction: A case report.

In addition to the esthetic outcomes, autologous breast reconstruction offers satisfactory functional results via sensory recovery of the flap. A herpes zoster infection developed after an autologous breast reconstruction provides objective evidence of spontaneous reinnervation in a reconstructed breast. One previous case of a herpes zoster infection on autologous latissimus dorsi flap has been reported to date; the infection developed 2 years after the breast reconstruction operation. However, our case presents a herpes zoster infection developing only 2 months after surgery. To our knowledge, the present case represents the first reported instance of a herpes zoster infection that developed shortly after the breast reconstruction using a latissimus dorsi flap.

Identifying cystogenic paracrine signaling molecules in cyst fluid of patients with polycystic kidney disease.

In autosomal dominant polycystic kidney disease (ADPKD) paracrine signaling molecules in cyst fluid can induce proliferation and cystogenesis of neighboring renal epithelial cells. However, the identity of this cyst-inducing factor is still unknown. The aim of this study was to identify paracrine signaling proteins in cyst fluid using a 3D in vitro cystogenesis assay. We collected cyst fluid from 15 ADPKD patients who underwent kidney or liver resection (55 cysts from 13 nephrectomies, 5 cysts from 2 liver resections). For each sample, the ability to induce proliferation and cyst formation was tested using the cystogenesis assay (RPTEC/TERT1 cells in Matrigel with cyst fluid added for 14 days). Kidney cyst fluid induced proliferation and cyst growth of renal epithelial cells in a dose-dependent fashion. Liver cyst fluid also induced cystogenesis. Using size exclusion chromatography, 56 cyst fluid fractions were obtained of which only the fractions between 30 and 100 kDa showed cystogenic potential. Mass spectrometry analysis of samples that tested positive or negative in the assay identified 43 candidate cystogenic proteins. Gene ontology analysis showed an enrichment for proteins classified as enzymes, immunity proteins, receptors, and signaling proteins. A number of these proteins have previously been implicated in ADPKD, including secreted frizzled-related protein 4, S100A8, osteopontin, and cysteine rich with EGF-like domains 1. In conclusion, both kidney and liver cyst fluids contain paracrine signaling molecules that drive cyst formation. Using size exclusion chromatography and mass spectrometry, we procured a candidate list for future studies. Ultimately, cystogenic paracrine signaling molecules may be targeted to abrogate cystogenesis in ADPKD.

GluA4 Dependent Plasticity Mechanisms Contribute to Developmental Synchronization of the CA3-CA1 Circuitry in the Hippocampus.

During the course of development, molecular mechanisms underlying activity-dependent synaptic plasticity change considerably. At immature CA3-CA1 synapses in the hippocampus, PKA-driven synaptic insertion of GluA4 AMPA receptors is the predominant mechanism for synaptic strengthening. However, the physiological significance of the developmentally restricted GluA4-dependent plasticity mechanisms is poorly understood. Here we have used microelectrode array (MEA) recordings in GluA4 deficient slice cultures to study the role of GluA4 in early development of the hippocampal circuit function. We find that during the first week in culture (DIV2-6) when GluA4 expression is restricted to pyramidal neurons, loss of GluA4 has no effect on the overall excitability of the immature network, but significantly impairs synchronization of the CA3 and CA1 neuronal populations. In the absence of GluA4, the temporal correlation of the population spiking activity between CA3-CA1 neurons was significantly lower as compared to wild-types at DIV6. Our data show that synapse-level defects in transmission and plasticity mechanisms are efficiently compensated for to normalize population firing rate at the immature hippocampal network. However, lack of the plasticity mechanisms typical for the immature synapses may perturb functional coupling between neuronal sub-populations, a defect frequently implicated in the context of developmentally originating neuropsychiatric disorders.

Depolarizing γ-aminobutyric acid contributes to glutamatergic network rewiring in epilepsy.

OBJECTIVE: Rewiring of excitatory glutamatergic neuronal circuits is a major abnormality in epilepsy. Besides the rewiring of excitatory circuits, an abnormal depolarizing γ-aminobutyric acidergic (GABAergic) drive has been hypothesized to participate in the epileptogenic processes. However, a remaining clinically relevant question is whether early post-status epilepticus (SE) evoked chloride dysregulation is important for the remodeling of aberrant glutamatergic neuronal circuits. METHODS: Osmotic minipumps were used to infuse intracerebrally a specific inhibitor of depolarizing GABAergic transmission as well as a functionally blocking antibody toward the pan-neurotrophin receptor p75 (p75NTR ). The compounds were infused between 2 and 5 days after pilocarpine-induced SE. Immunohistochemistry for NKCC1, KCC2, and ectopic recurrent mossy fiber (rMF) sprouting as well as telemetric electroencephalographic and electrophysiological recordings were performed at day 5 and 2 months post-SE. RESULTS: Blockade of NKCC1 after SE with the specific inhibitor bumetanide restored NKCC1 and KCC2 expression, normalized chloride homeostasis, and significantly reduced the glutamatergic rMF sprouting within the dentate gyrus. This mechanism partially involves p75NTR signaling, as bumetanide application reduced SE-induced p75NTR expression and functional blockade of p75NTR decreased rMF sprouting. The early transient (3 days) post-SE infusion of bumetanide reduced rMF sprouting and recurrent seizures in the chronic epileptic phase. INTERPRETATION: Our findings show that early post-SE abnormal depolarizing GABA and p75NTR signaling fosters a long-lasting rearrangement of glutamatergic network that contributes to the epileptogenic process. This finding defines promising and novel targets to constrain reactive glutamatergic network rewiring in adult epilepsy. Ann Neurol 2017;81:251-265.

Immunohistochemical study on roe deer haemal nodes.

BACKGROUND: The aim of this study was to investigate the immunohistochemical characteristics of the haemal nodes located in the abdominal and thoracic cavities in roe deer (Capreolus capreolus). MATERIALS AND METHODS: In this study, 2 adult male and 2 adult female roe deers in addition to 2 roe deer foetuses at the late foetal stages were used. The avidin-biotin-peroxidase technique was applied to anti-CD3, anti-CD79acy, anti--macrophage, anti-S100 primary antibodies. RESULTS: Positive results were gained for all the antibodies used. Many CD3 positive T-lymphocytes were seen in the lymphoid tissue areas of the foetal haemal nodes, in the germinal centre of the lymph follicles of the adult haemal nodes and in their lymphatic cords. CD79acy positive cells were less in number while S-100 protein was positive in both lymph follicles and in sinuses of the adult haemal nodes. In foetal haemal nodes, cells positive for S-100 protein were more prevalent in the sinuses. For the MAC387 primary antibody, there were positive macrophages seen in the capsule and trabeculae of the haemal nodes, located denser in males. Macrophage cells in the lymphoid structures and granulocytes in the sinuses of the haemal nodes of both adults and foetuses were positive for MAC387. CONCLUSIONS: This study is the first one to define the detailed structural features immunohistochemically in the haemal nodes of the roe deer. Even it's concluded that the results were similar with the other ruminants, it's also pointed out that there are certain differences among the foetal, adult male and female haemal nodes of the roe deer. (Folia Morphol 2018; 77, 2: 266-271).

First report on the nationwide incidence and prevalence of Type 1 diabetes among children in Turkey.

AIM: To report, for the first time, the incidence and prevalence of childhood Type 1 diabetes in Turkey using a nationwide registry. METHODS: Information on birth date, city of birth, diagnosis date and gender of all patients with Type 1 diabetes aged < 18 years were obtained from the Turkish Social Security Institute for the period from January 2011 to December 2013. RESULTS: There were 17 175 prevalent cases of Type 1 diabetes over the 3-year period. The prevalence of Type 1 diabetes was 0.75/1 000 (95% CI 0.74-0.76) and was higher in girls than in boys (0.79 vs 0.72 /1 000; P < 0.01). There were 2465 incident cases in 2013. The incidence was slightly higher among girls (50.6%) than boys (49.4%); the girl:boy case ratio was 1.02. The incidence was 10.4/100 000 for boys and 11.3/100 000 for girls. The age-standardized incidence rate was 10.8 per 100 000 (95% CI 10.1-11.5) according to the WHO standard population, estimated using the direct method. The mean patient age at diagnosis was 10.6 ± 4.6 years. The highest proportion of cases (40.6%) was diagnosed in children aged 10-14 years. CONCLUSIONS: This is the first study to report the incidence and prevalence of Type 1 diabetes in children in Turkey. The incidence of Type 1 diabetes reflects the geographical location of Turkey, bridging Asia and Europe, with the incidence being higher than in Asia but lower than in Europe.

Developmental switch in the kinase dependency of long-term potentiation depends on expression of GluA4 subunit-containing AMPA receptors.

The AMPA-receptor subunit GluA4 is expressed transiently in CA1 pyramidal neurons at the time synaptic connectivity is forming, but its physiological significance is unknown. Here we show that GluA4 expression is sufficient to alter the signaling requirements of long-term potentiation (LTP) and can fully explain the switch in the LTP kinase dependency from PKA to Ca2(+)/calmodulin-dependent protein kinase II during synapse maturation. At immature synapses, activation of PKA leads to a robust potentiation of AMPA-receptor function via the mobilization of GluA4. Analysis of GluA4-deficient mice indicates that this mechanism is critical for neonatal PKA-dependent LTP. Furthermore, lentiviral expression of GluA4 in CA1 neurons conferred a PKA-dependent synaptic potentiation and LTP regardless of the developmental stage. Thus, GluA4 defines the signaling requirements for LTP and silent synapse activation during a critical period of synapse development.

Highly Conserved Keratin-Associated Protein 7-1 Gene in Yak, Taurine and Zebu Cattle.

Keratin-associated proteins (KRTAPs) play a critical role in cross-linking the keratin intermediate filaments to build a hair shaft. The genetic polymorphisms of the bovine KRTAP7-1 gene were investigated for the first time in this study. The complete coding sequence of the KRTAP7-1 gene in 108 domestic yak, taurine and zebu cattle from China and Indonesia were successfully amplified using polymerase chain reaction and then directly sequenced. Only two single-nucleotide polymorphisms (one nonsynonymous at c.7C/G and another synonymous at c.21C/T) and three haplotypes (BOVIN-KRTAP7-1*A, B and C) were identified in the complete coding sequence of the bovine KRTAP7-1 gene among all animals. There was no polymorphism across three Chinese indigenous yak breeds and one Indonesian zebu cattle population, all sharing the BOVINKRTAP71*A haplotype. The four taurine cattle populations also had BOVIN-KRTAP7-1*A as the most common haplotype with a frequency of 0.80. The frequency of novel haplotype BOVIN-KRTAP7-1*B was only 0.07 present in one heterozygous animal in each of the four taurine cattle populations, while BOVINKRTAP7- 1*C was only found in a Simmental and a local Chinese Yellow cattle population with frequencies of 0.17 and 0.36, respectively. The monomorphic yak KRTAP7-1 gene in particular, and highly conserved bovine, sheep and goat KRTAP7-1 genes in general, demonstrated its unique intrinsic structural property (e.g., > 21% high glycine content) and primary functional importance in supporting the mechanical strength and shape of hair.

Effects of selenium on ischaemia-reperfusion injury in a rat testis model.

Selenium is shown to have beneficial effects on ischaemia-reperfusion (IR) injury. Our aim was to assess the effects of selenium on IR-induced testicular damage in terms of biochemical and histopathological evaluation. A total of 32 rats were randomised into four groups: control, IR, IR + selenium (IR + S) and S. Detorsion was applied after 3 h of torsion. Testicular tissue superoxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA), total antioxidant capacity (TAC) and DNA fragmentation levels were determined. Testicular tissue samples were examined by histopathological examination and terminal deoxynucleotidyl transferase dUTP nick end-labelling staining. The control, IR and IR + S groups had higher SOD values compared with the S group; SOD levels of the control and IR + S groups were higher than those of the IR group (P < 0.05). Further, MDA levels of the IR group were higher than those in the other three groups (P < 0.05). The IR group revealed lower TAC levels than the three groups (P < 0.05 for all). GSH levels of the IR group were significantly lower than those in the other three groups (P < 0.05 for all). In contrast, GSH levels of the IR + S group increased compared with those of the S group. The IR group had more DNA fragmentation than the control and S groups (P < 0.05). It is concluded that selenium possibly reduces oxidative stress and apoptosis caused by testicular IR injury in rats. The testicular protective effect of selenium appears to be mediated through its anti-apoptotic and antioxidative effects. However, selenium does not affect DNA fragmentation.

A Rare Case of Hanhart Syndrome with Mild Developmental Delay.

Hanhart Syndrome (OMIM 103300) is an extremely rare syndrome with some congenital malformations. It is characterized by hypoglossia, adactylia/hypodactylia, peromelia of arms and/or legs and micrognathia. The severity of the symptoms can differ from patient to patient. Some affected individuals may have only a part of these clinical features. In this case report, we want to present a Turkish girl with hypoglossia, micrognathia and peromelia who was diagnosed according to the clinical and radiographic findings.

Activity-dependent upregulation of presynaptic kainate receptors at immature CA3-CA1 synapses.

Presynaptic kainate-type glutamate receptors (KARs) regulate glutamate release probability and short-term plasticity in various areas of the brain. Here we show that long-term depression (LTD) in the area CA1 of neonatal rodent hippocampus is associated with an upregulation of tonic inhibitory KAR activity, which contributes to synaptic depression and causes a pronounced increase in short-term facilitation of transmission. This increased KAR function was mediated by high-affinity receptors and required activation of NMDA receptors, nitric oxide (NO) synthetase, and postsynaptic calcium signaling. In contrast, KAR activity was irreversibly downregulated in response to induction of long-term potentiation in a manner that depended on activation of the TrkB-receptor of BDNF. Both tonic KAR activity and its plasticity were restricted to early stages of synapse development and were lost in parallel with maturation of the network due to ongoing BDNF-TrkB signaling. These data show that presynaptic KARs are targets for activity-dependent modulation via diffusible messengers NO and BDNF, which enhance and depress tonic KAR activity at immature synapses, respectively. The plasticity of presynaptic KARs in the developing network allows nascent synapses to shape their response to incoming activity. In particular, upregulation of KAR function after LTD allows the synapse to preferentially pass high-frequency afferent activity. This can provide a potential rescue from synapse elimination by uncorrelated activity and also increase the computational dynamics of the developing CA3-CA1 circuitry.

Somatic STAT3 mutations in large granular lymphocytic leukemia.

BACKGROUND: T-cell large granular lymphocytic leukemia is a rare lymphoproliferative disorder characterized by the expansion of clonal CD3+CD8+ cytotoxic T lymphocytes (CTLs) and often associated with autoimmune disorders and immune-mediated cytopenias. METHODS: We used next-generation exome sequencing to identify somatic mutations in CTLs from an index patient with large granular lymphocytic leukemia. Targeted resequencing was performed in a well-characterized cohort of 76 patients with this disorder, characterized by clonal T-cell-receptor rearrangements and increased numbers of large granular lymphocytes. RESULTS: Mutations in the signal transducer and activator of transcription 3 gene (STAT3) were found in 31 of 77 patients (40%) with large granular lymphocytic leukemia. Among these 31 patients, recurrent mutational hot spots included Y640F in 13 (17%), D661V in 7 (9%), D661Y in 7 (9%), and N647I in 3 (4%). All mutations were located in exon 21, encoding the Src homology 2 (SH2) domain, which mediates the dimerization and activation of STAT protein. The amino acid changes resulted in a more hydrophobic protein surface and were associated with phosphorylation of STAT3 and its localization in the nucleus. In vitro functional studies showed that the Y640F and D661V mutations increased the transcriptional activity of STAT3. In the affected patients, downstream target genes of the STAT3 pathway (IFNGR2, BCL2L1, and JAK2) were up-regulated. Patients with STAT3 mutations presented more often with neutropenia and rheumatoid arthritis than did patients without these mutations. CONCLUSIONS: The SH2 dimerization and activation domain of STAT3 is frequently mutated in patients with large granular lymphocytic leukemia; these findings suggest that aberrant STAT3 signaling underlies the pathogenesis of this disease. (Funded by the Academy of Finland and others.).

Caffeine suppresses homologous recombination through interference with RAD51-mediated joint molecule formation.

Caffeine is a widely used inhibitor of the protein kinases that play a central role in the DNA damage response. We used chemical inhibitors and genetically deficient mouse embryonic stem cell lines to study the role of DNA damage response in stable integration of the transfected DNA and found that caffeine rapidly, efficiently and reversibly inhibited homologous integration of the transfected DNA as measured by several homologous recombination-mediated gene-targeting assays. Biochemical and structural biology experiments revealed that caffeine interfered with a pivotal step in homologous recombination, homologous joint molecule formation, through increasing interactions of the RAD51 nucleoprotein filament with non-homologous DNA. Our results suggest that recombination pathways dependent on extensive homology search are caffeine-sensitive and stress the importance of considering direct checkpoint-independent mechanisms in the interpretation of the effects of caffeine on DNA repair.