Monovalent Pseudo-Natural Product Degraders Supercharge the Native Degradation of IDO1 by KLHDC3.

Targeted protein degradation (TPD) modulates protein function beyond inhibition of enzyme activity or protein-protein interactions. Most degraders function by proximity induction, and directly bridge an E3 ligase with the target to be degraded. However, many proteins might not be addressable via proximity-based degraders, and other challenges, such as resistance acquisition, exist. Here, we identified pseudo-natural products derived from (-)-myrtanol, termed iDegs, that inhibit and induce degradation of the immunomodulatory enzyme indoleamine-2,3-dioxygenase 1 (IDO1) by a distinct mechanism. iDegs induce a unique conformational change and, thereby, boost IDO1 ubiquitination and degradation by the cullin-RING E3 ligase CRL2KLHDC3, which we identified to also mediate native IDO1 degradation. Therefore, iDegs supercharge the native proteolytic pathway of IDO1, rendering this mechanism of action distinct from traditional degrader approaches involving proteolysis-targeting chimeras (PROTACs) or molecular-glue degraders (MGDs). In contrast to clinically explored IDO1 inhibitors, iDegs reduce formation of kynurenine by both inhibition and induced degradation of the enzyme and should also modulate non-enzymatic functions of IDO1. This unique mechanism of action may open up new therapeutic opportunities for the treatment of cancer beyond classical inhibition of IDO1.

Discovery of Dihydro-1,4-Benzoxazine Carboxamides as Potent and Highly Selective Inhibitors of Sirtuin-1.

Sirtuins are signaling hubs orchestrating the cellular response to various stressors with roles in all major civilization diseases. Sirtuins remove acyl groups from lysine residues of proteins, thereby controlling their activity, turnover, and localization. The seven human sirtuins, SirT1-7, are closely related in structure, hindering the development of specific inhibitors. Screening 170,000 compounds, we identify and optimize SirT1-specific benzoxazine inhibitors, Sosbo, which rival the efficiency and surpass the selectivity of selisistat (EX527). The compounds inhibit the deacetylation of p53 in cultured cells, demonstrating their ability to permeate biological membranes. Kinetic analysis of inhibition and docking studies reveal that the inhibitors bind to a complex of SirT1 and nicotinamide adenine dinucleotide, similar to selisistat. These new SirT1 inhibitors are valuable alternatives to selisistat in biochemical and cell biological studies. Their greater selectivity may allow the development of better targeted drugs to combat SirT1 activity in diseases such as cancer, Huntington's chorea, or anorexia.

Enantioselective Synthesis of a Tricyclic, sp3 -Rich Diazatetradecanedione: an Amino Acid-Based Natural Product-Like Scaffold.

6-, 7-, and 8-membered rings are assembled from a linear precursor by successive cyclisation reactions to construct a tricyclic diazatricyclo[6.5.1.04, 9 ]-tetradecanedione scaffold. Advanced building blocks based on d-aspartic acid and l-pyroglutamic acid were combined by a sp3 -sp2 Negishi coupling. A carbamate-guided syn-diastereoselective epoxidation followed by an intramolecular epoxide opening allowed the construction of the piperidine ring. An efficient one-pot hydroxyl-group protection twofold deprotection reaction prepared the ground for the cyclisation to the bicycle. A final deprotection of the orthogonal protecting groups and lactamisation led to the novel, sp3 -rich tricycle. The final compound is a substrate mimic of peptidyl-prolyl cis-trans isomerases featuring a locked trans-amide bond. Cheminformatic analysis of 179 virtual derivatives indicates favourable physicochemical properties and drug-like characteristics. As proof of concept we, show a low micromolar activity in a fluorescence polarisation assay towards the FK506-binding protein 12.

Chemical Genetics Reveals a Role of dCTP Pyrophosphatase 1 in Wnt Signaling.

Cell-based screening is a powerful approach to identify novel chemical modulators and biological components of relevant biological processes. The canonical Wnt pathway is essential for normal embryonic development and tissue homeostasis, and its deregulation plays a crucial role in carcinogenesis. Therefore, the identification of new pathway members and regulators is of significant interest. By means of a cell-based assay monitoring Wnt signaling we identified the pyrrolocoumarin Pyrcoumin as inhibitor of canonical Wnt signaling. Target identification and validation revealed that Pyrcoumin is a competitive inhibitor of dCTP pyrophosphatase 1 (dCTPP1). We demonstrate a yet unknown interaction of dCTPP1 with ubiquitin carboxyl-terminal hydrolase (USP7) that is counteracted by dCTPP1 inhibitors. These findings indicate that dCTPP1 plays a role in regulation of Wnt/ß-catenin signaling most likely through a direct interaction with USP7.

Photoactivatable Rhodamine Spiroamides and Diazoketones Decorated with "Universal Hydrophilizer" or Hydroxyl Groups.

Photoactivatable rhodamine spiroamides and spirocyclic diazoketones emerged recently as synthetic markers applicable in multicolor super-resolution microscopy. However, their applicability in single molecule localization microscopy (SMLM) is often limited by aggregation, unspecific adhesion, and low reactivity caused by insufficient solubility and precipitation from aqueous solutions. We report here two synthetic modifications increasing the polarity of compact polycyclic and hydrophobic labels decorated with a reactive group: attachment of 3-sulfo-l-alanyl-beta-alanine dipeptide (a "universal hydrophilizer") or allylic hydroxylation in photosensitive rhodamine diazoketones (and spiroamides). The super-resolution images of tubulin and keratin filaments in fixed and living cells exemplify the performance of "blinking" spiroamides derived from N, N, N', N'-tetramethyl rhodamine.

Brief learning induces a memory bias for arousing-negative words: an fMRI study in high and low trait anxious persons.

Persons suffering from anxiety disorders display facilitated processing of arousing and negative stimuli, such as negative words. This memory bias is reflected in better recall and increased amygdala activity in response to such stimuli. However, individual learning histories were not considered in most studies, a concern that we meet here. Thirty-four female persons (half with high-, half with low trait anxiety) participated in a criterion-based associative word-learning paradigm, in which neutral pseudowords were paired with aversive or neutral pictures, which should lead to a valence change for the negatively paired pseudowords. After learning, pseudowords were tested with fMRI to investigate differential brain activation of the amygdala evoked by the newly acquired valence. Explicit and implicit memory was assessed directly after training and in three follow-ups at 4-day intervals. The behavioral results demonstrate that associative word-learning leads to an explicit (but no implicit) memory bias for negatively linked pseudowords, relative to neutral ones, which confirms earlier studies. Bilateral amygdala activation underlines the behavioral effect: Higher trait anxiety is correlated with stronger amygdala activation for negatively linked pseudowords than for neutrally linked ones. Most interestingly, this effect is also present for negatively paired pseudowords that participants could not remember well. Moreover, neutrally paired pseudowords evoked higher amygdala reactivity than completely novel ones in highly anxious persons, which can be taken as evidence for generalization. These findings demonstrate that few word-learning trials generate a memory bias for emotional stimuli, indexed both behaviorally and neurophysiologically. Importantly, the typical memory bias for emotional stimuli and the generalization to neutral ones is larger in high anxious persons.

Anticipating action effects with different attention foci is reflected in brain activation.

Anticipation is informed by experience. Having focused on action effects in the past will lead to differences when the focus is now on the effector. Boules-type throwing movements were presented as point-light displays of shoulder and arm-markers. Activation in motor-related areas measured with functional magnetic resonance imaging was compared between two tasks: Task A anticipating action effects and Task B judging the velocity of the hand marker. One group of participants performed a session of Task A followed by a session of Task B; the other group started with Task B followed by Task A. The group starting with Task A exhibited higher brain activation during Task A bilaterally in intraparietal areas and in right hemispheric frontal and premotor areas. These areas are known to be involved in effect estimation and action simulation. The second group showed higher activation during Task B in premotor cortex and human intraparietal area 3 of the right hemisphere. The results suggest that the instruction to focus on anticipating action effects facilitates the recruitment of core components of the simulation network during anticipation and when effect anticipation is not the primary intention.

Anticipating action effects recruits audiovisual movement representations in the ventral premotor cortex.

When table tennis players anticipate the course of the ball while preparing their motor responses, they not only observe their opponents striking the ball but also listen to events such as the sound of racket-ball contact. Because visual stimuli can be detected more easily when accompanied by a sound, we assumed that complementary sensory audiovisual information would influence the anticipation of biological motion, especially when the racket-ball contact is not presented visually, but has to be inferred from continuous movement kinematics and an abrupt sound. Twenty-six observers were examined with fMRI while watching point-light displays (PLDs) of an opposing table tennis player. Their task was to anticipate the resultant ball flight. The sound was presented complementary to the veracious event or at a deviant time point in its kinematics. Results showed that participants performed best in the complementary condition. Using a region-of-interest approach, fMRI data showed that complementary audiovisual stimulation elicited higher activation in the left temporo-occipital middle temporal gyrus (MTGto), the left primary motor cortex, and the right anterior intraparietal sulcus (aIPS). Both hemispheres also revealed higher activation in the ventral premotor cortex (vPMC) and the pars opercularis of the inferior frontal gyrus (BA 44). Ranking the behavioral effect of complementary versus conflicting audiovisual information over participants revealed an association between the complementary information and higher activation in the right vPMC. We conclude that the recruitment of movement representations in the auditory and visual modalities in the vPMC can be influenced by task-relevant cross-modal audiovisual interaction.

Enantioselective total synthesis of the lignan (+)-linoxepin.

An enantioselective total synthesis of the natural (+)-linoxepin (1) was accomplished in eleven steps from bromovanin (24). Key steps are a domino carbopalladation/ Mizoroki-Heck reaction with the formation of a pentacyclic system, an asymmetric hydroboration as well as an oxidative lactonization.

Stereoselective construction of the 5-hydroxy diazabicyclo[4.3.1]decane-2-one scaffold, a privileged motif for FK506-binding proteins.

A stereoselective synthesis of a derivatized bicyclic [4.3.1]decane scaffold based on an acyclic precursor is described. The key steps involve a Pd-catalyzed sp(3)-sp(2) Negishi-coupling, an asymmetric Shi epoxidation, and an intramolecular epoxide opening. Representative derivatives of this novel scaffold were synthesized and found to be potent inhibitors of the psychiatric risk factor FKBP51, which bound to FKBP51 with the intended molecular binding mode.

Timing accuracy in self-timed movements related to neural indicators of movement initiation.

Timely movement initiation is crucial in quick reactions or when a series of movements has to be strung together in a timed fashion to create a coordinated sequence. Stochastic neural variability can lead to misinitiation errors as reaction time studies suggest. Higher reaction times occur when preparatory neural activity reaches an initiation threshold later relative to shorter reaction times. Whether this also applies to self-timed movements is harder to scrutinize because they lack an external event that could serve as a reference for timing accuracy estimations. By example of a self-timed goal-oriented throwing task, we used a method that synchronizes the throwing movements by their kinematic profiles to assess relative timing differences in throwing release. We determined neural preparatory processes of the release using the movement-related electrophysiological Bereitschaftspotential (BP). By analyzing differences in shape and timing of the BP in delayed and non-delayed throws, two variables could be extracted that are related to timing differences on the kinematic level. First, temporal deviations in BP curves partly meet the kinematic deviations. Second, delayed releases were preceded by a short flattening of the BP curves prior to release. Thus, temporal and shape deviations in the neural movement initiation are assumed to delay self-timed movements.

Parietal and premotor cortices: activation reflects imitation accuracy during observation, delayed imitation and concurrent imitation.

This study investigated whether activation within areas belonging to the action observation and imitation network reveals a linear relation to the subsequent accuracy of imitating a bimanual rhythmic movement measured via a motion capturing system. 20 participants were scanned with functional magnetic resonance imaging (fMRI) when asked to imitate observed bimanual movements either concurrently versus with a delay (2s) or simply to observe the movements without imitation. Results showed that action observation relates to activation within classic mirror-related areas. Activation patterns were more widespread when participants were asked to imitate the movement. During observation with concurrent imitation, activation in the left inferior parietal lobe (IPL) was associated negatively with imitation accuracy. During observation in the delayed imitation condition, higher subsequent imitation accuracy was coupled with higher activation in the right superior parietal lobe (SPL) and the left parietal operculum (POp). During the delayed imitation itself, a negative association between imitation accuracy and brain activation was revealed in the right ventral premotor cortex (vPMC). We conclude that the IPL is involved in online comparison and visuospatial attention processes during imitation, the SPL provides a kinesthetic blueprint during movement observation, the POp preserves body identity, and the vPMC recruits motor representations--especially when no concurrent visual guidance is possible.

Prediction of human actions: expertise and task-related effects on neural activation of the action observation network.

The action observation network (AON) is supposed to play a crucial role when athletes anticipate the effect of others' actions in sports such as tennis. We used functional magnetic resonance imaging to explore whether motor expertise leads to a differential activation pattern within the AON during effect anticipation and whether spatial and motor anticipation tasks are associated with a differential activation pattern within the AON depending on participant expertise level. Expert (N=16) and novice (N=16) tennis players observed video clips depicting forehand strokes with the instruction to either indicate the predicted direction of ball flight (spatial anticipation) or to decide on an appropriate response to the observed action (motor anticipation). The experts performed better than novices on both tennis anticipation tasks, with the experts showing stronger neural activation in areas of the AON, namely, the superior parietal lobe, the intraparietal sulcus, the inferior frontal gyrus, and the cerebellum. When novices were contrasted with experts, motor anticipation resulted in stronger activation of the ventral premotor cortex, the supplementary motor area, and the superior parietal lobe than spatial anticipation task did. In experts, the comparison of motor and spatial anticipation revealed no increased activation. We suggest that the stronger activation of areas in the AON during the anticipation of action effects in experts reflects their use of the more fine-tuned motor representations they have acquired and improved during years of training. Furthermore, results suggest that the neural processing of different anticipation tasks depends on the expertise level.

Major depressive disorder alters perception of emotional body movements.

Much recent research has shown an association between mood disorders and an altered emotion perception. However, these studies were conducted mainly with stimuli such as faces. This is the first study to examine possible differences in how people with major depressive disorder (MDD) and healthy controls perceive emotions expressed via body movements. Thirty patients with MDD and thirty healthy controls observed the video scenes of human interactions conveyed by point-light displays (PLDs). They rated the depicted emotions and judged their confidence in their rating. Results showed that patients with MDD rated the depicted interactions more negatively than healthy controls. They also rated interactions with negative emotionality as being more intense and were more confident in their ratings. It is concluded that patients with MDD exhibit an altered emotion perception compared to healthy controls when rating emotions expressed via body movements depicted in PLDs.

Neural simulation of actions: effector- versus action-specific motor maps within the human premotor and posterior parietal area?

This study addresses the controversy over how motor maps are organized during action simulation by examining whether action simulation states, that is, motor imagery and action observation, run on either effector-specific and/or action-specific motor maps. Subjects had to observe or imagine three types of movements effected by the right hand or the right foot with different action goals. The functional magnetic resonance imaging results showed an action-specific organization within premotor and posterior parietal areas of both hemispheres during action simulation, especially during action observation. There were also less pronounced effector-specific activation sites during both simulation processes. It is concluded that the premotor and parietal areas contain multiple motor maps rather than a single, continuous map of the body. The forms of simulation (observation, imagery), the task contexts (movements related to an object, with usual/unusual effector), and the underlying reason for performing the simulation (rate your subjective success afterwards) lead to the specific use of different representational motor maps within both regions. In our experimental setting, action-specific maps are dominant especially, during action observation, whereas effector-specific maps are recruited to only a lesser degree.

How equivalent are the action execution, imagery, and observation of intransitive movements? Revisiting the concept of somatotopy during action simulation.

Jeannerod (2001) hypothesized that action execution, imagery, and observation are functionally equivalent. This led to the major prediction that these motor states are based on the same action-specific and even effector-specific motor representations. The present study examined whether hand and foot movements are represented in a somatotopic manner during action execution, imagery, and action observation. The experiment contained ten conditions: three execution conditions, three imagery conditions, three observation conditions, and one baseline condition. In the nine experimental conditions, participants had to execute, observe, or imagine right-hand extension/flexion movements or right-foot extension/flexion movements. The fMRI results showed a somatotopic organization within the contralateral premotor and primary motor cortex during motor imagery and motor execution. However, there was no clear somatotopic organization of action observation in the given regions of interest within the contralateral hemisphere, although observation of these movements activated these areas significantly.

Confidence in emotion perception in point-light displays varies with the ability to perceive own emotions.

One central issue in social cognitive neuroscience is that perceiving emotions in others relates to activating the same emotion in oneself. In this study we sought to examine how the ability to perceive own emotions assessed with the Toronto Alexithymia Scale related to both the ability to perceive emotions depicted in point-light displays and the confidence in these perceptions. Participants observed video scenes of human interactions, rated the depicted valence, and judged their confidence in this rating. Results showed that people with higher alexithymia scores were significantly less confident about their decisions, but did not differ from people with lower alexithymia scores in the valence of their ratings. Furthermore, no modulating effect of social context on the effect of higher alexithymia scores was found. It is concluded that the used stimuli are fit to investigate the kinematic aspect of emotion perception and possibly separate people with high and low alexithymia scores via confidence differences. However, a general difference in emotion perception was not detected in the present setting.

Motor familiarity: brain activation when watching kinematic displays of one's own movements.

The perception of action is influenced by the observer's familiarity with its movement. However, how does motor familiarity with own movement patterns modulate the visual perception of action effects? Cortical activation was examined with fMRI while 20 observers were watching videotaped point-light displays of markers on the shoulders, the right elbow, and wrist of an opposing table tennis player. The racket and ball were not displayed. Participants were asked to predict the invisible effect of the stroke, that is, the ball flight direction. Different table tennis models were used without the observers knowing and being informed in advance that some of the presented videos displayed their own movements from earlier training sessions. Prediction had to be made irrespective of the identity of the player represented by the four moving markers. Results showed that participants performed better when observing their "own" strokes. Using a region-of-interest approach, fMRI data showed that observing own videos was accompanied by stronger activation (compared to other videos) in the left angular gyrus of the inferior parietal lobe and the anterior rostral medial frontal cortex. Other videos elicited stronger activation than own videos in the left intraparietal sulcus and right supramarginal gyrus. We suggest that during action observation of motorically familiar movements, the compatibility between the observed action and the observers' motor representation is already coded in the parietal angular gyrus--in addition to the paracingulate gyrus. The activation in angular gyrus is presumably part of an action-specific effect retrieval that accompanies actor-specific prefrontal processing. The intraparietal sulcus seems to be sensitive to incongruence between observed kinematics and internal model representations, and this also influences processing in the supramarginal gyrus.

Transcriptional activation and production of tryptophan-derived secondary metabolites in arabidopsis roots contributes to the defense against the fungal vascular pathogen Verticillium longisporum.

The soil-borne fungal pathogen Verticillium longisporum causes vascular disease on Brassicaceae host plants such as oilseed rape. The fungus colonizes the root xylem and moves upwards to the foliage where disease symptoms become visible. Using Arabidopsis as a model for early gene induction, we performed root transcriptome analyses in response to hyphal growth immediately after spore germination and during penetration of the root cortex, respectively. Infected roots showed a rapid reprogramming of gene expression such as activation of transcription factors, stress-, and defense-related genes. Here, we focused on the highly coordinated gene induction resulting in the production of tryptophan-derived secondary metabolites. Previous studies in leaves showed that enzymes encoded by CYP81F2 and PEN2 (PENETRATION2) execute the formation of antifungal indole glucosinolate (IGS) metabolites. In Verticillium-infected roots, we found transcriptional activation of CYP81F2 and the PEN2 homolog PEL1 (PEN2-LIKE1), but no increase in antifungal IGS breakdown products. In contrast, indole-3-carboxylic acid (I3CA) and the phytoalexin camalexin accumulated in infected roots but only camalexin inhibited Verticillium growth in vitro. Whereas genetic disruption of the individual metabolic pathways leading to either camalexin or CYP81F2-dependent IGS metabolites did not alter Verticillium-induced disease symptoms, a cyp79b2 cyp79b3 mutant impaired in both branches resulted in significantly enhanced susceptibility. Hence, our data provide an insight into root-specific early defenses and suggest tryptophan-derived metabolites as active antifungal compounds against a vascular pathogen.