Architectures of Lipid Transport Systems for the Bacterial Outer Membrane.

How phospholipids are trafficked between the bacterial inner and outer membranes through the hydrophilic space of the periplasm is not known. We report that members of the mammalian cell entry (MCE) protein family form hexameric assemblies with a central channel capable of mediating lipid transport. The E. coli MCE protein, MlaD, forms a ring associated with an ABC transporter complex in the inner membrane. A soluble lipid-binding protein, MlaC, ferries lipids between MlaD and an outer membrane protein complex. In contrast, EM structures of two other E. coli MCE proteins show that YebT forms an elongated tube consisting of seven stacked MCE rings, and PqiB adopts a syringe-like architecture. Both YebT and PqiB create channels of sufficient length to span the periplasmic space. This work reveals diverse architectures of highly conserved protein-based channels implicated in the transport of lipids between the membranes of bacteria and some eukaryotic organelles.

Rapid Drink Challenge During High-resolution Manometry for Evaluation of Esophageal Emptying in Treated Achalasia.

BACKGROUND & AIMS: Incomplete esophageal emptying is a key variable predicting symptom relapse after achalasia treatment. Although optimally evaluated using the timed barium esophagogram (TBE), incomplete esophageal emptying can also be identified on rapid drink challenge (RDC) performed during high-resolution manometry. METHODS: We evaluated if RDC differentiates complete from incomplete esophageal emptying in treated patients with achalasia, against a TBE gold standard. Unselected treated patients with achalasia with both TBE (200 mL of low-density barium suspension) and RDC (200 mL of water in sitting position) were enrolled in 5 tertiary referral centers. TBE barium column height at 1, 2, and 5 minutes were compared with RDC variables: pressurizations >20 mmHg, maximal RDC pressurization, proportion of RDC time occupied by pressurizations, trans-esophagogastric junction gradient, and integrated relaxation pressure. RESULTS: Of 175 patients recruited (mean age, 59 years; 47% female), 138 (79%) were in clinical remission. Complete TBE emptying occurred in 45.1% at 1 minute, 64.0% at 2 minutes, and 73.1% at 5 minutes. RDC integrated relaxation pressure correlated strongly with TBE column height, and a 10-mmHg threshold discriminated complete from incomplete emptying at all 3 TBE time points with area under receiver operating characteristic curves of 0.85, 0.87, and 0.85, respectively. This threshold had high negative predictive values for complete emptying (88% at 2 minutes, 94% at 5 minutes), and modest positive predictive values for incomplete emptying (77% at 2 minutes, 62% at 5 minutes). CONCLUSIONS: RDC during high-resolution manometry is an effective surrogate for TBE in assessing esophageal emptying in treated patients with achalasia.

Improving the Diagnostic Yield of High-Resolution Esophageal Manometry for GERD: The "Straight Leg-Raise" International Study.

BACKGROUND & AIMS: The straight leg raise (SLR) maneuver during high-resolution manometry (HRM) can assess esophagogastric junction (EGJ) barrier function by measuring changes in intraesophageal pressure (IEP) when intra-abdominal pressure is increased. We aimed to determine whether increased esophageal pressure during SLR predicts pathologic esophageal acid exposure time (AET). METHODS: Adult patients with persistent gastroesophageal reflux disease (GERD) symptoms undergoing HRM and pH-impedance or wireless pH study off proton pump inhibitor were prospectively studied between July 2021 and March 2022. After the HRM Chicago 4.0 protocol, patients were requested to elevate 1 leg at 45º for 5 seconds while supine. The SLR maneuver was considered effective when intra-abdominal pressure increased by 50%. IEPs were recorded 5 cm above the lower esophageal sphincter at baseline and during SLR. GERD was defined as AET greater than 6%. RESULTS: The SLR was effective in 295 patients (81%), 115 (39%) of whom had an AET greater than 6%. Hiatal hernia (EGJ type 2 or 3) was seen in 135 (46%) patients. Compared with patients with an AET less than 6%, peak IEP during SLR was significantly higher in the GERD group (29.7 vs 13.9 mm Hg; P < .001). Using receiver operating characteristic analysis, an increase of 11 mm Hg of peak IEP from baseline during SLR was the optimal cut-off value to predict an AET greater than 6% (area under the receiver operating characteristic curve, 0.84; sensitivity, 79%; and specificity, 85%), regardless of the presence of hiatal hernia. On multivariable analysis, an IEP pressure increase during the SLR maneuver, EGJ contractile integral, EGJ subtype 2, and EGJ subtype 3, were found to be significant predictors of AET greater than 6% CONCLUSIONS: The SLR maneuver can predict abnormal an AET, thereby increasing the diagnostic value of HRM when GERD is suspected. CLINICALTRIALS: gov ID: NCT04813029.

Proximal Gastric Pressurization After Sleeve Gastrectomy Associates With Gastroesophageal Reflux.

INTRODUCTION: Sleeve gastrectomy (SG) results in persistent or de novo reflux more often than Roux-en-Y gastric bypass (RYGB). We investigated pressurization patterns in the proximal stomach on high-resolution manometry (HRM) to determine associations with reflux after SG. METHODS: Patients undergoing HRM and ambulatory pH-impedance monitoring after SG and RYGB over a 2-year period (2019-2020) were included. For each included patient, 2 symptomatic control patients with HRM and pH-impedance monitoring for reflux symptoms were identified within the same time frame; 15 asymptomatic healthy controls with HRM studies were also studied. Concurrent myotomy and preoperative diagnosis of obstructive motor disorders were exclusions. Conventional HRM metrics, esophagogastric junction (EGJ) pressures, contractile integral (EGJ-CI), acid exposure time (AET), and reflux episode numbers were extracted. Intragastric pressure was sampled at baseline, during swallows, and with straight leg raise maneuver, and compared with intraesophageal pressure and reflux burden. RESULTS: Patient cohorts included 36 SG patients, 23 RYGB patients, 113 symptomatic controls, and 15 asymptomatic controls. While both SG and RYGB patients pressurized the stomach during swallows and straight leg raise, SG patients had higher AET (median 6.0% vs 0.2%), reflux episode numbers (median 63.0 vs 37.5), and baseline intragastric pressure (median 17.3 mm Hg vs 13.1 mm Hg) ( P < 0.001). SG patients also had lower trans-EGJ pressure gradients when reflux episodes were >80 or AET was >6.0% ( P = 0.018 and 0.08, respectively, compared with no pathologic reflux). On multivariable analysis, SG status and low EGJ-CI independently associated with AET and reflux episode numbers ( P ≤ 0.04). DISCUSSION: Impaired EGJ barrier function and proximal gastric pressurization after SG are associated with gastroesophageal reflux, especially during strain maneuvers.

Racial and Ethnic Disparities in Germline Genetic Testing of Patients With Young-Onset Colorectal Cancer.

BACKGROUND & AIMS: Up to 20% of younger patients (age <50 years) diagnosed with colorectal cancer (CRC) have germline mutations in cancer susceptibility genes. Germline genetic testing may guide clinical management and facilitate earlier intervention in affected relatives. Few studies have characterized differences in genetic testing by race/ethnicity. METHODS: We identified young adults (age 18-49 years) diagnosed with CRC between 2009 and 2017 in 2 health systems in Dallas, TX. We evaluated referral to genetic counseling, attendance at genetic counseling appointments, and receipt of germline genetic testing by race/ethnicity. RESULTS: Of 385 patients with young-onset CRC (median age at diagnosis 44.4 years), 176 (45.7%) were Hispanic, 98 (25.4%) non-Hispanic Black, and 111 (28.8%) non-Hispanic White. Most patients (76.9%) received immunohistochemistry (IHC) for mismatch repair proteins, and there was no difference in receipt of IHC by race/ethnicity. However, a lower proportion of Black patients were referred to genetic counseling (50.0% vs White patients 54.1% vs Hispanic patients 65.9%; P = .02) and attended genetic counseling appointments (61.2% vs 81.7% White patients vs 86.2% Hispanic patients; P < .01). Of 141 patients receiving genetic testing, 38 (27.0%) had a pathogenic or likely pathogenic variant in a cancer susceptibility gene. An additional 33 patients (23.4%) had variants of uncertain significance, of which 84.8% occurred in racial/ethnic minorities. CONCLUSIONS: In a diverse population of patients diagnosed with young-onset CRC, we observed racial/ethnic differences in referral to and receipt of germline genetic testing. Our findings underscore the importance of universal genetic testing to address racial/ethnic disparities in young-onset CRC.

Is Colorectal Cancer in Patients Younger Than 50 Years of Age the Same Disease as in Older Patients?

The incidence of colorectal cancer (CRC) has declined steadily in persons over age 50 years of age, largely due to screening. In contrast, incidence rates have increased rapidly in younger adults (<50 years of age),1 raising the question of whether young-onset CRC is a distinct disease with unique biologic features or if it is the same disease occurring at a younger age. Studies comparing younger and older patients diagnosed with CRC have reported differences in clinical and molecular features, including tumor location, stage, and histology.2-6 However, increasing use of screening colonoscopy in the population has changed characteristics of CRC diagnosed over 50 years of age (eg, higher proportion now diagnosed with proximal tumors and/or at an earlier stage),7 so it is challenging to draw conclusions about the importance of these findings. To address this challenge, we compared characteristics of younger and older CRC patients in a predominantly unscreened population.

Ursodeoxycholic Acid Decreases Incidence of Primary Biliary Cholangitis and Biliary Complications After Liver Transplantation: A Meta-Analysis.

After liver transplantation (LT), the role of ursodeoxycholic acid (UDCA) is not well characterized. We examine the effect of UDCA after LT in the prophylaxis of biliary complications (BCs) in all-comers for LT and the prevention of recurrent primary biliary cholangitis (rPBC) in patients transplanted for PBC. Two authors searched PubMed/MEDLINE and Embase from January 1990 through December 2018 to identify all studies that evaluate the effectiveness of UDCA prophylaxis after LT for BCs in all LT recipients and rPBC after LT in patients transplanted for PBC. Odds ratios (ORs) were calculated for endpoints of the BC study. Pooled recurrence rates were calculated for rPBC. The study was conducted in accordance with the Preferred Reporting Items for Systematic Review and Meta-Analysis guidelines. A total of 15 studies were included, comprising 530 patients in the analysis for BCs and 1727 patients in the analysis for rPBC. UDCA was associated with decreased odds of BCs (OR, 0.70; 95% confidence interval [CI], 0.52-0.93; P = 0.01) and biliary stones and sludge (OR, 0.49; 95% CI, 0.24-0.77; P = 0.004). Prophylactic use of UDCA did not affect the odds of biliary stricture. For patients transplanted for PBC, the rate of rPBC was lower with the prophylactic use of UDCA (IR 16.7%; 95% CI, 0.114%-22.0%; I2 = 36.1%) compared with not using prophylactic UDCA (IR 23.1%; 95% CI, 16.9%-29.3%; I2 = 86.7%). UDCA after LT reduces the odds of BC and bile stones and sludge in all-comer LT recipients and reduces or delays the incidence of rPBC in patients transplanted for PBC. UDCA use after LT could be considered in all LT recipients to reduce the odds of BC and may be particularly beneficial for patients transplanted for PBC by reducing the incidence of rPBC.

Recreational marijuana use is not associated with worse outcomes after renal transplantation.

As marijuana (MJ) legalization is increasing, kidney transplant programs must develop listing criteria for marijuana users. However, no data exist on the effect of MJ on kidney allograft outcomes, and there is no consensus on whether MJ use should be a contraindication to transplantation. We retrospectively reviewed 1225 kidney recipients from 2008 to 2013. Marijuana use was defined by positive urine toxicology screen and/or self-reported recent use. The primary outcome was death at 1 year or graft failure (defined as GFR<20 mL/min/1.73 m2 ). The secondary outcome was graft function at 1 year. Using logistic regression analyses, we compared these outcomes between MJ users and non-users. Marijuana use was not associated with worse primary outcomes by unadjusted (odds ratio 1.07, 95% CI 0.45-2.57, P=.87) or adjusted (odds ratio 0.79, 95% CI 0.28-2.28, P=.67) analysis. Ninety-two percent of grafts functioned at 1 year. Among these, the mean creatinine (1.52, 95% CI 1.39-1.69 vs 1.46, 95% CI 1.42-1.49; P=.38) and MDRD GFR (50.7, 95% CI 45.6-56.5 vs 49.5, 95% CI 48.3-50.7; P=.65) were similar between groups. Isolated recreational MJ use is not associated with poorer patient or kidney allograft outcomes at 1 year. Therefore, recreational MJ use should not necessarily be considered a contraindication to kidney transplantation.

The Milan score: A novel manometric tool for a more efficient diagnosis of gastro-esophageal reflux disease.

OBJECTIVE: A definitive diagnosis of gastroesophageal reflux disease (GERD) depends on endoscopic and/or pH-study criteria. However, high resolution manometry (HRM) can identify factors predicting GERD, such as ineffective esophageal motility (IEM), esophago-gastric junction contractile integral (EGJ-CI), evaluating esophagogastric junction (EGJ) type and straight leg raise (SLR) maneuver response. We aimed to build and externally validate a manometric score (Milan Score) to stratify the risk and severity of the disease in patients undergoing HRM for suspected GERD. METHODS: A population of 295 consecutive patients undergoing HRM and pH-study for persistent typical or atypical GERD symptoms was prospectively enrolled to build a model and a nomogram that provides a risk score for AET > 6%. Collected HRM data included IEM, EGJ-CI, EGJ type and SLR. A supplemental cohort of patients undergoing HRM and pH-study was also prospectively enrolled in 13 high-volume esophageal function laboratories across the world in order to validate the model. Discrimination and calibration were used to assess model's accuracy. Gastroesophageal reflux disease was defined as acid exposure time >6%. RESULTS: Out of the analyzed variables, SLR response and EGJ subtype 3 had the highest impact on the score (odd ratio 18.20 and 3.87, respectively). The external validation cohort consisted of 233 patients. In the validation model, the corrected Harrel c-index was 0.90. The model-fitting optimism adjusted calibration slope was 0.93 and the integrated calibration index was 0.07, indicating good calibration. CONCLUSIONS: A novel HRM score for GERD diagnosis has been created and validated. The MS might be a useful screening tool to stratify the risk and the severity of GERD, allowing a more comprehensive pathophysiologic assessment of the anti-reflux barrier. TRIAL REGISTRATION: ClinicalTrials.gov (Identifier: NCT05851482).

In situ cryo-electron tomography reveals the asymmetric architecture of mammalian sperm axonemes.

The flagella of mammalian sperm display non-planar, asymmetric beating, in contrast to the planar, symmetric beating of flagella from sea urchin sperm and unicellular organisms. The molecular basis of this difference is unclear. Here, we perform in situ cryo-electron tomography of mouse and human sperm, providing the highest-resolution structural information to date. Our subtomogram averages reveal mammalian sperm-specific protein complexes within the microtubules, the radial spokes and nexin-dynein regulatory complexes. The locations and structures of these complexes suggest potential roles in enhancing the mechanical strength of mammalian sperm axonemes and regulating dynein-based axonemal bending. Intriguingly, we find that each of the nine outer microtubule doublets is decorated with a distinct combination of sperm-specific complexes. We propose that this asymmetric distribution of proteins differentially regulates the sliding of each microtubule doublet and may underlie the asymmetric beating of mammalian sperm.

Automated tracing of microtubules in electron tomograms of plastic embedded samples of Caenorhabditis elegans embryos.

The ability to rapidly assess microtubule number in 3D image stacks from electron tomograms is essential for collecting statistically meaningful data sets. Here we implement microtubule tracing using 3D template matching. We evaluate our results by comparing the automatically traced centerlines to manual tracings in a large number of electron tomograms of the centrosome of the early Caenorhabditis elegans embryo. Furthermore, we give a qualitative description of the tracing results for three other types of samples. For dual-axis tomograms, the automatic tracing yields 4% false negatives and 8% false positives on average. For single-axis tomograms, the accuracy of tracing is lower (16% false negatives and 14% false positives) due to the missing wedge in electron tomography. We also implemented an editor specifically designed for correcting the automatic tracing. Besides, this editor can be used for annotating microtubules. The automatic tracing together with a manual correction significantly reduces the amount of manual labor for tracing microtubule centerlines so that large-scale analysis of microtubule network properties becomes feasible.

Solid bolus swallows during high-resolution manometry complement multiple rapid swallows in predicting symptoms following antireflux surgery.

BACKGROUND: Solid bolus swallows (SBS) assess esophageal peristalsis on high-resolution manometry (HRM). It remains unknown if increase in contraction vigor (contraction reserve) following SBS (SBS-CR) provides information similar to that observed following multiple rapid swallows (MRS-CR). METHODS: Clinical and HRM data from adult patients (n = 96, age 58.2 years, 61.5% female) undergoing esophageal testing prior to anti-reflux surgery (ARS) were analyzed if both MRS and SBS were available, and compared to similar data from healthy asymptomatic volunteers (n = 18, age 27.5 years, 55.6% female). Patients reported esophageal symptoms before and after ARS using 5-point Likert scales evaluating symptom frequency and severity; scores for each patient were averaged to determine global symptoms. Distal contractile integral (DCI) from single swallows was compared to MRS DCI and SBS DCI; the ratio of single swallow DCI to MRS DCI or SBS DCI ≥1 determined the presence of contraction reserve. KEY RESULTS: Multiple rapid swallows CR was concordant with SBS CR in 55.6% of healthy volunteers, 100% of absent contractility, 100% of ineffective esophageal motility (IEM) and 56.9% without IEM. Correlation between MRS DCI and SBS DCI was highest when IEM criteria were met (Spearman's rho 0.998). While neither MRS CR nor SBS CR presence differed by presenting symptoms or global symptom score at baseline, post-ARS global symptom burden was highest in the absence of both MRS CR and SBS CR, especially in patients with esophageal hypomotility using Chicago Classification version 4.0 criteria (p = 0.03). CONCLUSIONS & INFERENCES: Multiple rapid swallows and SBS are complementary in predicting post-ARS symptom burden.

AreTomo: An integrated software package for automated marker-free, motion-corrected cryo-electron tomographic alignment and reconstruction.

AreTomo, an abbreviation for Alignment and Reconstruction for Electron Tomography, is a GPU accelerated software package that fully automates motion-corrected marker-free tomographic alignment and reconstruction in a single package. By correcting in-plane rotation, translation, and importantly, the local motion resulting from beam-induced motion from tilt to tilt, AreTomo can produce tomograms with sufficient accuracy to be directly used for subtomogram averaging. Another major application is the on-the-fly reconstruction of tomograms in parallel with tilt series collection to provide users with real-time feedback of sample quality allowing users to make any necessary adjustments of collection parameters. Here, the multiple alignment algorithms implemented in AreTomo are described and the local motions measured on a typical tilt series are analyzed. The residual local motion after correction for global motion was found in the range of ± 80 Å, indicating that the accurate correction of local motion is critical for high-resolution cryo-electron tomography (cryoET).

The elegans of spindle assembly.

The Caenorhabditis elegans one-cell embryo is a powerful system in which to study microtubule organization because this large cell assembles both meiotic and mitotic spindles within the same cytoplasm over the course of 1 h in a stereotypical manner. The fertilized oocyte assembles two consecutive acentrosomal meiotic spindles that function to reduce the replicated maternal diploid set of chromosomes to a single-copy haploid set. The resulting maternal DNA then unites with the paternal DNA to form a zygotic diploid complement, around which a centrosome-based mitotic spindle forms. The early C. elegans embryo is amenable to live-cell imaging and electron tomography, permitting a detailed structural comparison of the meiotic and mitotic modes of spindle assembly.

Electron cryotomography of intact motile cilia defines the basal body to axoneme transition.

Cells use motile cilia to generate force in the extracellular space. The structure of a cilium can be classified into three subdomains: the intracellular basal body (BB) that templates cilium formation, the extracellular axoneme that generates force, and the transition zone (TZ) that bridges them. While the BB is composed of triplet microtubules (TMTs), the axoneme is composed of doublet microtubules (DMTs), meaning the cilium must convert between different microtubule geometries. Here, we performed electron cryotomography to define this conversion, and our reconstructions reveal identifying structural features of the BB, TZ, and axoneme. Each region is distinct in terms of microtubule number and geometry, microtubule inner proteins, and microtubule linkers. TMT to DMT conversion occurs within the BB, and microtubule geometry changes to axonemal by the end of the TZ, followed by the addition of axoneme-specific components essential for cilium motility. Our results provide the highest-resolution images of the motile cilium to date and reveal how BBs template axonemes.

Insights into centriole geometry revealed by cryotomography of doublet and triplet centrioles.

Centrioles are cylindrical assemblies comprised of 9 singlet, doublet, or triplet microtubules, essential for the formation of motile and sensory cilia. While the structure of the cilium is being defined at increasing resolution, centriolar structure remains poorly understood. Here, we used electron cryo-tomography to determine the structure of mammalian (triplet) and Drosophila (doublet) centrioles. Mammalian centrioles have two distinct domains: a 200 nm proximal core region connected by A-C linkers, and a distal domain where the C-tubule is incomplete and a pair of novel linkages stabilize the assembly producing a geometry more closely resembling the ciliary axoneme. Drosophila centrioles resemble the mammalian core, but with their doublet microtubules linked through the A tubules. The commonality of core-region length, and the abrupt transition in mammalian centrioles, suggests a conserved length-setting mechanism. The unexpected linker diversity suggests how unique centriolar architectures arise in different tissues and organisms.

The role of γ-tubulin in centrosomal microtubule organization.

As part of a multi-subunit ring complex, γ-tubulin has been shown to promote microtubule nucleation both in vitro and in vivo, and the structural properties of the complex suggest that it also seals the minus ends of the polymers with a conical cap. Cells depleted of γ-tubulin, however, still display many microtubules that participate in mitotic spindle assembly, suggesting that γ-tubulin is not absolutely required for microtubule nucleation in vivo, and raising questions about the function of the minus end cap. Here, we assessed the role of γ-tubulin in centrosomal microtubule organisation using three-dimensional reconstructions of γ-tubulin-depleted C. elegans embryos. We found that microtubule minus-end capping and the PCM component SPD-5 are both essential for the proper placement of microtubules in the centrosome. Our results further suggest that γ-tubulin and SPD-5 limit microtubule polymerization within the centrosome core, and we propose a model for how abnormal microtubule organization at the centrosome could indirectly affect centriole structure and daughter centriole replication.

Hierarchical assembly of the eggshell and permeability barrier in C. elegans.

In metazoans, fertilization triggers the assembly of an extracellular coat that constitutes the interface between the embryo and its environment. In nematodes, this coat is the eggshell, which provides mechanical rigidity, prevents polyspermy, and is impermeable to small molecules. Using immunoelectron microscopy, we found that the Caenorhabditis elegans eggshell was composed of an outer vitelline layer, a middle chitin layer, and an inner layer containing chondroitin proteoglycans. The switch between the chitin and proteoglycan layers was achieved by internalization of chitin synthase coincident with exocytosis of proteoglycan-containing cortical granules. Inner layer assembly did not make the zygote impermeable as previously proposed. Instead, correlative light and electron microscopy demonstrated that the permeability barrier was a distinct envelope that formed in a separate step that required fatty acid synthesis, the sugar-modifying enzyme PERM-1, and the acyl chain transfer enzyme DGTR-1. These findings delineate the hierarchy of eggshell assembly and define key molecular mechanisms at each step.

Centrosome size sets mitotic spindle length in Caenorhabditis elegans embryos.

Just as the size of an organism is carefully controlled, the size of intracellular structures must also be regulated. The mitotic spindle is a supramolecular machine that generates the forces which separate sister chromatids during mitosis. Although spindles show little size variation between cells of the same type, spindle length can vary at least 10-fold between different species. Recent experiments on spindle length showed that in embryonic systems spindle length varied with blastomere size. Furthermore, a comparison between two Xenopus species showed that spindle length was dependent on some cytoplasmic factor. These data point toward mechanisms to scale spindle length with cell size. Centrosomes play an important role in organizing microtubules during spindle assembly. Here we use Caenorhabditis elegans to study the role of centrosomes in setting spindle length. We show that spindle length correlates with centrosome size through development and that a reduction of centrosome size by molecular perturbation reduces spindle length. By systematically analyzing centrosome proteins, we show that spindle length does not depend on microtubule density at centrosomes. Rather, our data suggest that centrosome size sets mitotic spindle length by controlling the length scale of a TPXL-1 gradient along spindle microtubules.

Identification of a peptide fragment of DSCR1 that competitively inhibits calcineurin activity in vitro and in vivo.

Calcineurin phosphatase activity regulates the nuclear localization of the nuclear factor of activated T cells (NFAT) family of transcription factors during immune challenge. Calcineurin inhibitors, such as the cyclosporin A-cyclophilin A and FK506-FKBP12 complexes, regulate this enzymatic activity noncompetitively by binding at a site distinct from the enzyme active site. A family of endogenous protein inhibitors of calcineurin was recently identified and shown to block calcineurin-mediated NFAT nuclear localization and transcriptional activation. One such inhibitor, Down Syndrome Critical Region 1 (DSCR1), functions in T cell activation, cardiac hypertrophy, and angiogenesis. We have identified a small region of DSCR1 that is a potent inhibitor of calcineurin activity in vitro and in vivo.