Analysis of rare disruptive germline mutations in 2135 enriched BRCA-negative breast cancers excludes additional high-impact susceptibility genes.

BACKGROUND: Breast cancer has a significant heritable basis, of which ∼60% remains unexplained. Testing for BRCA1/BRCA2 offers useful discrimination of breast cancer risk within families, and identification of additional breast cancer susceptibility genes could offer clinical utility. PATIENTS AND METHODS: We included 2135 invasive breast cancer cases recruited via the Breast and Ovarian Cancer Susceptibility study, a retrospective UK study of familial breast cancer. ELIGIBILITY CRITERIA: female, BRCA-negative, white European ethnicity, and one of: (i) breast cancer family history, (ii) bilateral disease, (iii) young age of onset (<30 years), and (iv) concomitant ovarian cancer. We undertook exome sequencing of cases and carried out gene-level burden testing of rare damaging variants against those from 51 377 ethnicity-matched population controls from gnomAD. RESULTS: 159/2135 (7.4%) cases had a qualifying variant in an established breast cancer susceptibility gene, with minimal evidence of signal in other cancer susceptibility genes. Known breast cancer susceptibility genes PALB2, CHEK2, and ATM were the only genes to retain statistical significance after correcting for multiple testing. Due to the enrichment of hereditary cases in the series, we had good power (>80%) to detect a gene of BRCA1-like risk [odds ratio (OR) = 10.6] down to a population minor allele frequency of 4.6 × 10-5 (1 in 10 799, less than one-tenth that of BRCA1)and of PALB2-like risk (OR = 5.0) down to a population minor allele frequency of 2.8 × 10-4 (1 in 1779, less than half that of PALB2). Power was lower for identification of novel moderate penetrance genes (OR = 2-3) like CHEK2 and ATM. CONCLUSIONS: This is the largest case-control whole-exome analysis of enriched breast cancer published to date. Whilst additional breast cancer susceptibility genes likely exist, those of high penetrance are likely to be of very low mutational frequency. Contention exists regarding the clinical utility of such genes.

What is the value of simulated patient assessment in structured clinical examinations of undergraduate students, and how should it be used?

INTRODUCTION: The aim of this study was to explore correlations between clinical assessor and simulated patient (SP) scores drawn from summative Integrated Structured Clinical Examination (ISCE) and inform the best use of SP scores in future assessments. MATERIALS AND METHODS: This retrospective study explores summative clinical assessor and formative SP numeric scores drawn from summative ISCE assessments spanning three academic years (2017-18 to 2019-20). Analyses were carried out using R 3.5.1 (R Core Team, 2018), with the stats package. RESULTS: The sample consisted of 169 final-year BDS students across the three cohorts and included 95 females (56.2%) and 74 males (43.8%). Data from eight substations where SPs were included, were explored. Kendall's Tau, a non-parametric correlation, was used to investigate the relationships between the assessor and SP scores. Clinical assessor scores were out of a total of 20 points across various assessed domains within each substation. The formative SP assessment was out of 10 points with the same five affective domains related to communication included in each substation. Overall, the assessor and patient substation scores were not correlated (τ = 0.04, p = .272) indicating that communication skills alone, as assessed by patients, do not correlate with more holistic performance across other domains. There was significant positive correlation for two of the eight substations with the other substations showing very little correlation. CONCLUSION: This study shows that assessment of student performance by SPs does not show a correlation with examiner scores and may provide additional information relating to affective skills of students. Notwithstanding the limitations of this study, the findings underscore the need to investigate further the value of involvement of SPs in clinical assessments to explore if scores by SPs can be used to enhance the validity of assessments if used summatively.

Reflections on learning and enhancing communication skills through community engagement: a student perspective.

Students in Peninsula School of Dentistry (PSD), Plymouth, undertake community engagement projects during the first two years of their undergraduate curriculum. These projects involve interaction with a variety of specific community groups and the planning and delivery of an appropriate and meaningful oral health intervention. Many of the project outcomes are based on enhancing communication skills and encouraging students to transfer these into their patient treatment sessions. This report draws on the experience of students who undertook two specific projects to demonstrate how they feel this is achieved.

Slow platelet recovery after PBPC transplantation from unrelated donors.

The effects of the composition of PBPC grafts from matched related donors (MRDs) and matched unrelated donors (MUDs) have not been compared. In a single-center study, the compositions of 55 MRD PBPC grafts and 33 MUD grafts were studied for their effect on the rate of engraftment in patients who had evidence of donor cell engraftment on day +28. The MUD grafts came more frequently from young male donors and contained more CD34(+) cells but similar numbers of colony-forming units granulocyte-macrophage (CFU-GM) and burst forming units-erythroid. The recovery of neutrophils to >500/mm(3) was equally fast in both groups, but recovery of platelets to >20,000/mm(3) was significantly delayed in the MUD group (P<0.001). The MUD group also required more transfusions of platelets and red cells. Patients receiving grafts containing low numbers of CFU-GM had markedly delayed platelet recovery. The patients with the slowest engraftment tended to have prolonged transportation times. Storage experiments suggested a major loss of viable CD34(+) cells and CFU-GM when undiluted PBPC products are stored at room temperature. The data suggest that a fraction of the MUD grafts suffer during transportation. In vitro proliferation assays should be part of the validation and auditing of transportation of MUD grafts.

Comparative Efficacy of Rabeprazole and Pantoprazole in the Control of Nocturnal Acid Output and Intragastric Acidity

BACKGROUND/AIMS: Nocturnal reflux is a largely undiagnosed and unmanaged condition predisposing to multiple esophageal complications. We evaluated the effects of rabeprazole and pantoprazole on nocturnal intragastric pH and gastric acid output during Day 1 of therapy following the consumption of standard meals. METHODS: The study had a double-blinded, randomized, two-way crossover design, and involved 15 patients with a history of mild reflux. Following an overnight fast, patients were given either rabeprazole (20 mg) or pantoprazole (40 mg) prior to the first of three standard Western meals. They then underwent overnight continuous intragastric pH monitoring and gastric acid output measurement. The drug effect was analyzed using a two-treatment, two-period crossover mixed model. RESULTS: The percentage of time during which the mean intragastric pH was greater than 4.0 and gastric acid output was less than 2.0 was higher for oral rabeprazole (p<0.05). The inhibition of acid output was greater for rabeprazole at almost all time points. Furthermore, the mean time-matched pH values differed significantly over the first 8.3 hours (p<0.05). CONCLUSIONS: On day 1, oral rabeprazole inhibited acid output to a greater extent and for a longer period than pantoprazole, and the intragastric pH was significantly higher for rabeprazole than for pantoprazole over the first 8.3 hours.

Requirement of Nck adaptors for actin dynamics and cell migration stimulated by platelet-derived growth factor B.

The Nck family of Src homology (SH) 2/SH3 domain adaptors functions to link tyrosine phosphorylation induced by extracellular signals with downstream regulators of actin dynamics. We investigated the role of mammalian Nck adaptors in signaling from the activated platelet-derived growth factor (PDGF) receptor (PDGFbetaR) to the actin cytoskeleton. We report here that Nck adaptors are required for cytoskeletal reorganization and chemotaxis stimulated by PDGF-B. Analysis of tyrosine-phosphorylated proteins demonstrated that Crk-associated substrate (p130(Cas)), not the activated PDGFbetaR itself, is the major Nck SH2 domain-binding protein in PDGF-B-stimulated cells. Both Nck- and p130(Cas)-deficient cells fail to display cytoskeletal rearrangements, including the formation of membrane ruffles and the disassembly of actin bundles, typically shown by their WT counterparts in response to PDGF-B. Furthermore, Nck and p130(Cas) colocalize in phosphotyrosine-enriched membrane ruffles induced by PDGF-B in NIH 3T3 cells. These results suggest that Nck adaptors play an essential role in linking the activated PDGFbetaR with actin dynamics through a pathway that involves p130(Cas).

Impaired PBPC collection in patients with myeloma after high-dose melphalan.

BACKGROUND: Tandem stem cell transplantation is an important treatment option for patients with myeloma and some additional tumors. In an attempt to reduce the contamination of the stem cell graft with tumor cells, patients with myeloma who entered complete remission after the first transplant underwent a second episode of mobilization to obtain progenitor cells for the second transplant. METHODS: Twenty-two patients with myeloma participated in the study. The first mobilization utilized CY, etoposide and filgrastim. The second mobilization used the same regimen, but seven patients received only filgrastim. The interval between the two collection periods was 6 months (median; range 4-9 months). The preparative regimen for the first transplant consisted of melphalan 200 mg/m(2). RESULTS: The number of total white cells collected during the two collection episodes was similar: 10.8+/-1.6 x 10(8)/kg white cells vs. 11.8+/-1.7 x 10(8)/kg white cells (P=0.63). The collected CD34(+) cell dose was much larger during the first collection: 45.2+/-8.4 x 10(6)/kg vs. 6.9+/-2.7 x 10(6)/kg (P<0.001). Similarly, the collected colony-forming unit (CFU)-GM dose was much larger during the first collection: 295.4+/-59.3 x 10(4)/kg vs. 67.3+/-21.6x10(4)/kg (P<0.001). While the CD34(+) cells collected during the two collection episodes correlated significantly (r=0.55, P<0.01); the first dose was a median of 14.9-fold larger. DISCUSSION: No laboratory parameter was able reliably to predict the results of the second collection. A second mobilization/collection episode as part of a tandem transplant approach carries a considerable risk of failing to obtain sufficient progenitor cells.

Mechanisms of CAS substrate domain tyrosine phosphorylation by FAK and Src.

Tyrosine phosphorylation of CAS (Crk-associated substrate, p130(Cas)) has been implicated as a key signaling step in integrin control of normal cellular behaviors, including motility, proliferation, and survival. Aberrant CAS tyrosine phosphorylation may contribute to cell transformation by certain oncoproteins, including v-Crk and v-Src, and to tumor growth and metastasis. The CAS substrate domain (SD) contains 15 Tyr-X-X-Pro motifs, which are thought to represent the major tyrosine phosphorylation sites and to function by recruiting downstream signaling effectors, including c-Crk and Nck. CAS makes multiple interactions, direct and indirect, with the tyrosine kinases Src and focal adhesion kinase (FAK), and as a result of this complexity, several plausible models have been proposed for the mechanism of CAS-SD phosphorylation. The objective of this study was to provide experimental tests of these models in order to determine the most likely mechanism(s) of CAS-SD tyrosine phosphorylation by FAK and Src. In vitro kinase assays indicated that FAK has a very poor capacity to phosphorylate CAS-SD, relative to Src. However, FAK expression along with Src was found to be important for achieving high levels of CAS tyrosine phosphorylation in COS-7 cells, as well as recovery of CAS-associated Src activity toward the SD. Structure-functional studies for both FAK and CAS further indicated that FAK plays a major role in regulating CAS-SD phosphorylation by acting as a docking or scaffolding protein to recruit Src to phosphorylate CAS, while a secondary FAK-independent mechanism involves Src directly bound to the CAS Src-binding domain (SBD). Our results do not support models in which FAK either phosphorylates CAS-SD directly or phosphorylates CAS-SBD to promote Src binding to this site.

Focal adhesion kinase is involved in mechanosensing during fibroblast migration.

Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase localized at focal adhesions and is believed to mediate adhesion-stimulated effects. Although ablation of FAK impairs cell movement, it is not clear whether FAK might be involved in the guidance of cell migration, a role consistent with its putative regulatory function. We have transfected FAK-null fibroblasts with FAK gene under the control of the tetracycline repression system. Cells were cultured on flexible polyacrylamide substrates for the detection of traction forces and the application of mechanical stimulation. Compared with control cells expressing wild-type FAK, FAK-null cells showed a decrease in migration speed and directional persistence. In addition, whereas FAK-expressing cells responded to exerted forces by reorienting their movements and forming prominent focal adhesions, FAK-null cells failed to show such responses. Furthermore, FAK-null cells showed impaired responses to decreases in substrate flexibility, which causes control cells to generate weaker traction forces and migrate away from soft substrates. Cells expressing Y397F FAK, which cannot be phosphorylated at a key tyrosine site, showed similar defects in migration pattern and force-induced reorientation as did FAK-null cells. However, other aspects of F397-FAK cells, including the responses to substrate flexibility and the amplification of focal adhesions upon mechanical stimulation, were similar to that of control cells. Our results suggest that FAK plays an important role in the response of migrating cells to mechanical input. In addition, phosphorylation at Tyr-397 is required for some, but not all, of the functions of FAK in cell migration.

The tubulogenic activity associated with an activated form of Flt-1 kinase is dependent on focal adhesion kinase.

Focal adhesion kinase (FAK) is known to be located at the intersection between extracellular matrix and growth factor signaling pathways to regulate cell motility. We have shown previously that an activated form (BCR-FLTm1) of Flt-1 kinase, a receptor for vascular endothelial growth factor, had a tubulogenic activity not only in endothelial cells but also in fibroblastic cells. Here we show that tubulogenesis by BCR-FLTm1 depends on FAK and that FAK tyrosine phosphorylation and association with an activated Flt-1 receptor complex is increased after vascular endothelial growth factor stimulation of NIH3T3 cells overexpressing Flt-1.

Spiral computed tomography for the diagnosis of pulmonary embolism in critically ill surgical patients: a comparison with pulmonary angiography.

HYPOTHESIS: Spiral computed tomographic pulmonary angiography (CTPA) is sensitive and specific in diagnosing pulmonary embolism (PE) in critically ill surgical patients. DESIGN: Prospective study comparing CTPA with the criterion standard, pulmonary angiography (PA). SETTING: Surgical intensive care unit of an academic hospital. PATIENTS: Twenty-two critically ill surgical patients with clinical suspicion of PE. The CTPAs and PAs were independently read by 4 radiologists (2 for each test) blinded to each other's interpretation. Clinical suspicion was classified as high, intermediate, or low according to predetermined criteria. All but 2 patients had marked pulmonary parenchymal disease at the time of the event that triggered evaluation for PE. INTERVENTIONS: Computed tomographic pulmonary angiography and PA in 22 patients, venous duplex scan in 19. RESULTS: Eleven patients (50%) had evidence of PE on PA, 5 in central and 6 in peripheral pulmonary arteries. The sensitivity and specificity of CTPA was, respectively, 45% and 82% for all PEs, 60% and 100% for central PEs, and 33% and 82% for peripheral PEs. Duplex scanning was 40% sensitive and 100% specific in diagnosing PE. The independent reviewers disagreed only in 14% of CTPA and 14% of PA interpretations. There were no differences in risk factors or clinical characteristics between patients with and without PE. The level of clinical suspicion was identical in the 2 groups. CONCLUSIONS: Pulmonary angiography remains the gold standard for the diagnosis of PE in critically ill surgical patients. Computed tomographic pulmonary angiography needs further evaluation in this population.

Age-based outcomes of cheilectomy for the treatment of hallux rigidus.

The results of cheilectomy, performed on 67 consecutive patients with hallux rigidus resulting in primary extraarticular symptoms are presented. Four patients who underwent subsequent fusion were rated as failures. Follow-up evaluation, averaging 65 months (28-117) on 53 additional patients available for follow-up, revealed an average AOFAS score of 80. with 91% of the patients stating that they were currently better than before surgery. There was a statistically significant higher mean score (89) in patients over 60 years of age at the time of surgery. There were no differences between other age groups, preoperative grade, duration of symptoms, or length of follow-up. Cheilectomy should be the treatment of choice for hallux rigidus with predominantly extra-articular symptoms, especially in patients over 60 years of age.

Angiographic embolization of bilateral internal iliac arteries to control life-threatening hemorrhage after blunt trauma to the pelvis.

Angiographic embolization of bleeding pelvic vessels is increasingly used in patients with pelvic injuries. Temporary angiographic embolization of bilateral internal iliac arteries (TAEBIIA) is occasionally necessary. From November 1991 to March 1998, 30 consecutive patients (mean age of 43 years, mean Injury Severity Score of 25) with complex pelvic fractures underwent TAEBIIA to control severe hemorrhage not responding to subselective embolization. Angiography revealed multiple sources of pelvic bleeding in 28 (93%) patients. In the two remaining patients, no bleeding was identified but TAEBIIA was done empirically. Thirteen patients had laparotomies before TAEBIIA with unsuccessful bleeding control, and the remaining 17 had TAEBIIA as the primary treatment. After TAEBIIA 90 per cent of patients had successful clinical (27 of 30) and radiographic (25 of 28) control of bleeding. Of the three patients who continued to bleed after TAEBIIA two were successfully re-embolized and one died of acute cardiac failure before any further intervention was attempted. TAEBIIA had a success rate of 97 per cent (29 of 30) in controlling pelvic hemorrhage without significant complications related to it. TAEBIIA is a safe and effective alternative to subselective embolization in controlling retroperitoneal bleeding in selected patients with blunt pelvic trauma.

Vena cavography with CO(2) versus with iodinated contrast material for inferior vena cava filter placement: a prospective evaluation.

PURPOSE: To determine whether carbon dioxide (CO(2)) vena cavography can safely guide the placement of inferior vena cava (IVC) filters. MATERIALS AND METHODS: One hundred nineteen patients were prospectively enrolled in this study. CO(2 )cavograms were obtained and evaluated for IVC diameter, location of renal veins, and presence of thrombus and venous anomalies. If CO(2 )cavography was judged to be adequate, an IVC filter was deployed. After filter placement, cavography was performed with iodinated contrast material; these images were compared with the CO(2) cavograms. RESULTS: Two patients experienced mild side effects related to venous CO(2) injection. Comparison of cavograms obtained with CO(2) and iodinated contrast-enhanced material showed the caval size to be within 3 mm in all 119 patients. In 116 patients (97.5%), CO(2) cavography was judged to be adequate, and in 115 patients, filters were placed. In three (2.5%) patients, it was necessary to perform iodinated contrast-enhanced cavography before filter deployment. All six cases of venous anomaly and 11 (78.6%) of 14 cases of thrombosis were clearly identified with CO(2) cavography. One filter was maldeployed owing to misinterpretation of the CO(2) cavogram. CONCLUSION: CO(2) cavography is well tolerated, safe, and adequate for identification of the parameters necessary for filter deployment. It is especially valuable in patients with a history of reaction to iodinated contrast material or renal insufficiency.

The acute effects of postoperative radiation therapy on the transverse rectus abdominis myocutaneous flap used in immediate breast reconstruction.

PURPOSE: To analyze the acute effects of postoperative radiation therapy on the transverse rectus abdominis myocutaneous (TRAM) flap reconstruction following modified radical mastectomy for breast cancer. METHODS AND MATERIALS: Twenty-five consecutive patients were treated with postoperative radiation therapy after TRAM flap reconstruction between 1985 and 1999. The radiation records for these patients were retrospectively reviewed. Information regarding treatment techniques, timing, and dose was obtained and correlated with the extent of erythema, desquamation, and the need for treatment break. RESULTS: The median age was 48 years. The median dose of chest wall radiation was 5040 cGy. Additional boost doses were delivered in 13 patients. Twelve patients (48%) developed mild erythema in the treatment field during the course of treatment and 13 patients (52%) developed moderate (40%) or brisk (12%) erythema. Only 10 patients (40%) developed any kind of desquamation; 5 patients (20%) developed dry desquamation and another 5 patients (20%) developed moist desquamation. No patients required a break in the course of treatment because of acute side effects. None of the parameters evaluated (the use of chemotherapy prior to radiation, the interval between surgery and radiation, smoking, prior incidence of fat necrosis, the use of bolus during radiation, and the use of a boost) were predictive of an increased incidence of either the extent of erythema or the development of desquamation in the treatment field. CONCLUSION: Postmastectomy radiation for TRAM flap reconstruction is well tolerated and is not associated with an increased incidence of acute side effects. Radiation technique and the use of preradiation chemotherapy do not appear to be correlated with an increased incidence of acute side effects.

Anti-apoptotic role of focal adhesion kinase (FAK). Induction of inhibitor-of-apoptosis proteins and apoptosis suppression by the overexpression of FAK in a human leukemic cell line, HL-60.

Focal adhesion kinase (FAK) has an anti-apoptotic role in anchorage-dependent cells via an unknown mechanism. To elucidate the role of FAK in anti-apoptosis, we have established several FAK cDNA-transfected HL-60 cell lines and examined whether FAK-transfected cells have resistance to apoptotic stimuli. FAK-transfected HL-60 (HL-60/FAK) cells were highly resistant to apoptosis induced with hydrogen peroxide (1 mm) and etoposide (50 microg/ml) compared with the parental HL-60 cells or the vector-transfected cells, when determined using viability assay, DNA fragmentation, and flow cytometry analysis. Because no proteolytic cleavage of pro-caspase 3 to mature caspase 3 fragment was observed in HL-60/FAK cells, FAK was presumed to inhibit an upstream signal pathway leading to the activation of caspase 3. HL-60/FAK activated the phosphatidylinositide 3'-OH-kinase-Akt survival pathway and exhibited significant activation of NF-kappaB with marked induction of inhibitor-of-apoptosis proteins (IAPs: cIAP-1, cIAP-2, XIAP), regardless of the hydrogen peroxide-treated or untreated conditions, whereas no significant IAPs were detected in the parental or vector-transfected HL-60 cells. Apoptotic agents induced higher NF-kappaB activation in HL-60/FAK cells than in HL-60/Vect cells, and it appeared that sustained NF-kappaB activation is critical to the anti-apoptotic states in HL-60/FAK cells. Mutagenesis of FAK cDNA revealed that Y397 and Y925, which are involved in the tyrosine-phosphorylation sites, were prerequisite for the anti-apoptotic activity as well as induction of IAPs, and that K454, which is involved in the kinase activity, was also required for the full anti-apoptotic activity of FAK. Taken together, we have demonstrated definitively that FAK-transfected HL-60 cells, otherwise sensitive to apoptosis, become resistant to the apoptotic stimuli. We conclude that FAK activates the phosphatidylinositide 3'-OH-kinase-Akt survival pathway with the concomitant activation of NF-kB and induction of IAPs, which ultimately inhibit apoptosis by inhibiting caspase-3 cascade.

The use of colorflow duplex scanning to detect significant renal artery stenosis.

The purpose of this study was to review our institutional experience with colorflow duplex scanning in detecting significant renal artery stenosis and to validate the criteria used: renal artery peak systolic velocity (PSV) >/=200 cm/sec and renal-to-aortic peak systolic ratio (RAR) >/=3.5. The results of renal artery duplex and arteriography in 58 patients (107 kidneys) who underwent both exams were reviewed. Arteriography revealed 32 main renal arteries with >/=60% stenosis. The PSV criterion detected 29, for a sensitivity of 91%, specificity of 75%, positive predictive value (PPV) of 60%, negative predictive value (NPV) of 95%, and accuracy of 79%. Using RAR >/=3.5 provided a sensitivity of 72%, specificity of 92%, PPV of 79%, NPV of 88%, and accuracy of 86%. In a subset of 36 kidneys that had hilar scans, the criteria of acceleration time (AT) >/=100 cm/sec and index (AI)

Angiographic embolization for intraperitoneal and retroperitoneal injuries.

Angiographic embolization (AE) has been used extensively for bleeding control after injuries to the face and neck. Its role in abdominal trauma requires further exploration. We reviewed the medical records of 137 consecutive patients who underwent angiography with the intent to embolize bleeding sites within the abdomen. Of them, 97 (71%) had blunt and 40 (29%) had penetrating trauma. AE was performed for hemorrhage associated with pelvic fractures (97 patients), liver lacerations (n = 26), renal lacerations (n = 12), splenic lacerations (n = 5), other injuries (n = 9), and multiple injuries (n = 12). On angiography, 102 patients were found to have bleeding sites and underwent AE, with angiographic and clinical bleeding control in 93 (91%). The rate of successful hemostasis by AE was identical in blunt and penetrating trauma patients. There was no major morbidity after AE. No factors predicted patients with a high likelihood to have a positive angiogram. Patients who had AE before or after a period of attempted hemodynamic stabilization in the intensive care unit were no different with respect to hemodynamic parameters immediately before AE or effectiveness of AE for bleeding control. AE is a safe and effective method for controlling bleeding after blunt and penetrating intra- and retroperitoneal injuries. Early AE may be used in selected patients as a front-line therapeutic intervention that offers expeditious hemostasis and prevents delays in definitive bleeding control.

Crk-associated substrate p130(Cas) interacts with nephrocystin and both proteins localize to cell-cell contacts of polarized epithelial cells.

Crk-associated substrate (p130(Cas), Cas) is a docking protein first recognized as having elevated phosphotyrosine content in mammalian cells transformed by v-Src and v-Crk oncoproteins. Subsequent studies have implicated Cas in the control of normal cell behavior through its roles in integrin-mediated signal transduction and organization of the actin cytoskeleton at sites of cell adhesion. In this study, we sought to gain new insight into normal Cas function by identifying previously unrecognized interacting proteins. A yeast two-hybrid screen using the C-terminal region of Cas as a bait identified the Src homology 3 (SH3) domain of the mouse "nephrocystin" protein-orthologous to a human protein whose loss of function leads to the cystic kidney disease familial juvenile nephronophthisis. The putative full-length mouse and partial canine nephrocystin sequences were deduced from cDNA clones. Additional studies using epitope-tagged mouse nephrocystin indicated that nephrocystin and Cas can interact in mammalian cells and revealed that both proteins prominently localize at or near sites of cell-cell contact in polarized Madin-Darby canine kidney epithelial cells. Our findings provide novel insight into the normal cellular activities regulated by both Cas and nephrocystin, and raise the possibility that these proteins have a related function in polarized epithelial cells.