Motor neurons with limb-innervating character in the cervical spinal cord are sculpted by apoptosis based on the Hox code in chick embryo.

In the developing chick embryo, a certain population of motor neurons (MNs) in the non-limb-innervating cervical spinal cord undergoes apoptosis between embryonic days 4 and 5. However, the characteristics of these apoptotic MNs remain undefined. Here, by examining the spatiotemporal profiles of apoptosis and MN subtype marker expression in normal or apoptosis-inhibited chick embryos, we found that this apoptotic population is distinguishable by Foxp1 expression. When apoptosis was inhibited, the Foxp1+ MNs survived and showed characteristics of lateral motor column (LMC) neurons, which are of a limb-innervating subtype, suggesting that cervical Foxp1+ MNs are the rostral continuation of the LMC. Knockdown and misexpression of Foxp1 did not affect apoptosis progression, but revealed the role of Foxp1 in conferring LMC identity on the cervical MNs. Furthermore, ectopic expression of Hox genes that are normally expressed in the brachial region prevented apoptosis, and directed Foxp1+ MNs to LMC neurons at the cervical level. These results indicate that apoptosis in the cervical spinal cord plays a role in sculpting Foxp1+ MNs committed to LMC neurons, depending on the Hox expression pattern.

Significance of p53-Binding Protein 1 Nuclear Foci in Cervical Squamous Intraepithelial Lesions: Association With High-Risk Human Papillomavirus Infection and P16INK4a Expression.

As p53-binding protein 1 (53BP1) localizes to the sites of DNA double-strand breaks and rapidly forms nuclear foci (NF), and its presence may be an indicator of endogenous genomic instability (GIN). We previously showed that 53BP1 NF in cervical cells increase with neoplastic progression, indicating the significance of 53BP1 expression for the estimation of malignant potential during cervical carcinogenesis. This study aimed to further elucidate the impact of 53BP1 expression as a biomarker for cervical squamous intraepithelial lesion (SIL). A total of 81 tissue samples, including 17 of normal cervical epithelium, 22 of cervical intraepithelial neoplasia (CIN) 1, 21 of CIN2, and 21 of CIN3, from patients positive for high-risk human papillomavirus (HR-HPV) were used for double-label immunofluorescence of 53BP1 and Ki-67/p16INK4a expression and HR-HPV in situ hybridization. We analyzed associations between 53BP1 expression type with parameters such as CIN grade, HR-HPV infection status, p16INK4a expression, and CIN prognosis. Expression type of 53BP1 was significantly associated with histological grade of CIN and HR-HPV in situ hybridization signal pattern (P < .0001). There was a significant correlation between 53BP1 and p16INK4a expression levels (r = .73, P < .0001). However, there was no association between 53BP1 expression type and CIN prognosis. We propose that 53BP1 expression type is a valuable biomarker for SIL, which can help estimate the grade and GIN of cervical lesions reflecting replication stress caused by the integration of HR-HPV to the host genome.

Malignant transformation from mature cystic teratoma of the ovary.

We present a case of malignant change of an ovarian mature cystic teratoma. Our patient was a 48-year-old woman and she visited a primary care doctor presenting with abdominal pain. At her first visit, her pelvic tumor measured 70 × 50 mm by ultrasonography. She was diagnosed as rupture of the malignant tumor occurred secondary to mature cystic teratoma and she took the surgery (abdominal total hysterectomy, bilateral oophorectomy and partial omentectomy). Pathologic diagnosis was squamous cell carcinoma (SCC) occurred secondary to mature cystic teratoma. Treatment with paclitaxel/carboplatin (TC chemotherapy) and gemcitabine hydrochloride/carboplatin (GC chemotherapy) after operation was not effective, and the refractory ileus resulting from rapid progression of the disease continued. She was died of disease progression 7 months after the diagnosis of ovarian cancer. We discuss about the clinical characteristics of malignant transformation of mature cystic teratoma and considered about the treatment of the ovarian SCC.

Improvement in the prognosis of ovarian cancer in the era before addition of molecular targeting therapy.

OBJECTIVE: The prognosis of ovarian cancer has improved because of platinum- and taxane-containing chemotherapy. We investigated the 5-year disease-specific overall survival and prognostic factors of patients with advanced ovarian cancer to elucidate the change in clinical course of ovarian cancer with the advance of chemotherapy for patients who developed relapse in the era before the addition of molecular targeting therapy. METHODS: We reviewed the clinical course of 134 patients with advanced ovarian cancer (FIGO Stage III and IV) treated in the past 11 years (1999-2010). We classified the patients into two groups: those who had been diagnosed with ovarian cancer from 1999 to 2005 (Group A) and those who had been diagnosed from 2006 to 2010 (Group B). We compared the 5-year disease-specific overall survival and median survival rates between these two groups. We also investigated the prognostic factors of 104 patients who developed relapse. RESULTS: The 5-year disease-specific overall survival rate was significantly higher in Group B than A (67.0% vs. 38.6%; P = 0.032). Chemotherapy containing pegylated liposomal doxorubicin hydrochloride, non-clear cell adenocarcinoma and intestinal resection were independent prognostic factors. CONCLUSIONS: The induction of new chemotherapeutic drugs and the increased variation of second- or third-line chemotherapy affected the improvement in overall survival of patients with advanced epithelial ovarian cancer.

Immune thrombocytopenia associated with solid cancer.

We describe a case of immune thrombocytopenia (ITP) associated with ovarian cancer. At the patient's first visit to hospital, high platelet-associated IgG and low platelet count (74 × 10(9)/L) were noted on blood test. She was diagnosed as having ITP complicated by ovarian cancer. Four days after surgery, the platelet count had increased to within the normal range. This is the first report of a patient with ITP complicated by ovarian cancer in which the platelet count reverted to normal soon after surgery for the ovarian cancer. We also investigated the characteristics of similar solid cancers with ITP at National Kyushu Cancer Center, Fukuoka, Japan.

[Nogitecan Hydrochloride Treatment for Patients with Recurrent Ovarian Cancer].

Nogitecan hydrochloride(topotecan)has shown efficacy in patients with recurrent ovarian cancer, but has not been used widely in Japan. We evaluated the efficacy and adverse effects of topotecan in 12 patients (median age, 62 years) treated for recurrent ovarian cancer between 2000 and 2013. Four patients had relapsed after primary treatment, and 8 had relapsed at least twice. Seven patients had been treated with more than 3 prior regimens. Initial treatment of the 12 patients consisted of intravenous topotecan (1.0-1.4 mg/m2/day) for 5 consecutive days. Initial doses were based on previous chemotherapy and/ or renal function, with reduced doses administered to patients with severe adverse effects during prior courses of treatment. The 12 patients received a total of 54 courses of topotecan(range, 1-15 courses). Of these 12 patients, one achieved a partial response and 6 had stable disease. The median time to progression was 14.4 weeks. All 12 patients had grade 3-4 myelosuppression, while none had febrile neutropenia or severe non-hematologic toxicities. Patients who received higher doses or increased courses of chemotherapy had apparently more severe adverse events. These findings suggested that topotecan should be used as a second- or third-line treatment, rather than later, in patients with tumor recurrence, with its dose reduced according to the physical status of each patient. Such strategies may enhance both the efficacy and safety of topotecan in patients with recurrent ovarian cancer.

Secondary leukemia after chemotherapy and/or radiotherapy for gynecologic neoplasia.

OBJECTIVE: Secondary leukemia is a known complication of chemotherapy and radiotherapy. It was generally recognized that leukemia secondary to chemotherapy was due to the use of alkylating agents in the treatment of ovarian cancer. Recently, many types of chemotherapeutic agents have been used in the treatment of gynecologic malignancies in addition to ovarian cancer. We analyzed the clinical characteristics and outcome of patients with recent onset of secondary leukemia after the treatment of gynecologic cancer to consider the diagnosis and management of secondary leukemia. MATERIALS AND METHODS: We reviewed the clinical charts and follow-up data of patients with gynecologic malignancies treated in the past 20 years. During this period, 2482 newly diagnosed invasive gynecologic cancers were treated in our institution. All patients with secondary leukemia were analyzed for clinical background, latency period (interval between the diagnosis of primary carcinoma and the development of leukemia), treatment, and outcome. We also reviewed the literature for secondary leukemia under gynecology using the PubMed. RESULTS: Four patients were found to have developed secondary leukemia after the treatment of gynecologic malignancies during this period. The cumulative risk of secondary leukemia was approximately 0.38%. All patients received platinum-based chemotherapy. Two patients received combination chemotherapy and/or bone marrow transplantation, and 1 of these 2 patients lived more than 6 years but died of recurrent ovarian cancer. CONCLUSIONS: Long survival time might be expected in patients who show complete response to bone marrow transplantation and/or combination chemotherapy for secondary leukemia. In recent years, we have aggressively used various types of anticancer drugs for the treatment of not only ovarian cancer but also uterine cervical cancer and endometrial cancer. Physicians need to keep in mind the risk of secondary leukemia in the follow-up of long-term survivors after several courses of chemotherapy and radiotherapy.

Prediction of histological types of endometrial cancer by endometrial cytology.

AIM: Few studies have examined the accuracy of preoperative endometrial cytology in diagnosing low- and high-risk histology in women with endometrial cancer (EC). This single-institutional retrospective study compared the accuracy of endometrial cytology and biopsy in preoperatively predicting low-risk and high-risk histology of EC. METHODS: Between January 2006 and March 2013, 198 women with EC were examined by endometrial cytology, endometrial biopsy and hysterectomy specimen in National Kyushu Cancer Center. Among these women, 110 had endometrial cytology samples available to compare with endometrial biopsy, and were enrolled in our study (mean age ± standard deviation: 59.57 ± 10.32 years). Single-use plastic endometrial suction curettes were used in 12 of the 110 cases and thin metallic curettes for the rest. RESULTS: For type 2 EC, which includes grade 3 endometrioid adenocarcinoma and non-endometrioid histology, biopsy was 67.6% sensitive (25/37) and 84.9% specific (62/73); whereas cytology was 70.3% sensitive (26/37) and 91.8% specific (67/73). Cytology precisely diagnosed only one of 14 cases of serous carcinoma, but it diagnosed 11 of the 14 cases as type 2 EC, and its accuracy in distinguishing EC types was not inferior to endometrial biopsy (10/14). For EC, 9.1% (10/110) were unevaluable using biopsy, significantly more than the 0% (0/110) by cytology (P = 0.002). CONCLUSION: Although preoperative prediction of serous carcinoma was difficult, endometrial cytology had a higher evaluable rate for EC types. Endometrial cytology may complement endometrial biopsy in preoperative women with EC.

Neurogenin2 expression together with NeuroM regulates GDNF family neurotrophic factor receptor α1 (GFRα1) expression in the embryonic spinal cord.

In many regions of the nervous system, the combinatorial action of transcriptional factors specifies the individual fate of neuronal subtypes. Contrary to this, we report that a single transcriptional factor controls a phenotype shared by different subtypes of neurons, namely the expression of a neurotrophic factor receptor in the spinal cord. Along the dorsoventral axis of the chick embryo spinal cord, the expression pattern of a specific receptor for glial cell line derived-neurotrophic factor (GDNF family of receptors α1: GFRα1) was related to that of two basic helix-loop-helix (bHLH) transcriptional factors (NeuroM and Neurogenin2: Ngn2). In ovo electroporation in the chick embryo revealed that the overexpression of NeuroM alone was sufficient to induce ectopic GFRα1 expression without overt neuronal differentiation, whereas the suppression of NeuroM activity resulted in the specific loss of GFRα1 expression, indicating that NeuroM may act as a differentiation factor for GFRα1 expression. Ngn2 overexpression was also sufficient to induce precocious GFRα1 expression. However, the forced expression of both obligate suppressor and activator forms of Ngn2 also induced aberrant GFRα1 expression. Thus, any deviation from an optimum level of Ngn2 expression resulted in aberrant GFRα1 expression. Consistent with this, manipulation of Ngn2 expression levels by other bHLH factors also resulted in ectopic GFRα1 expression. For example, the downregulation by Ascl1 and the upregulation by Ptf1a induced ectopic GFRα1 expression, irrespective of endogenous expression patterns of Ascl1 and Ptf1a (Ascl1/Ptf1) in the spinal cord. The suppression of Ascl1/Ptf1a activities abolished Ngn2 and GFRα1 expression, even in Ascl1/Ptf1a-negative regions. These data indicate the presence of a distinct regulatory sequence for a determinant of GFRα1 expression, in which Ascl1/Ptf1a may competitively intervene to stochastically modulate default Ngn2 expression levels. Thus, Ngn2 together with NeuroM serves as readout to regulate GFRα1 expression, which occurs in multiple subtypes of spinal neurons.

Dysgerminoma of the Left Ovary in a Patient with Balanced Translocation 46X, t(X:1) (q22;q21): A Case Report.

We report a case of dysgerminoma in a 22-year-old woman diagnosed with chromosomal abnormality, balanced translocation 46X,t(X:1)(q22;q21). She had received hormone replacement therapy for 7 years for primary amenorrhea. She visited a primary care physician because of lower abdominal distension, and a large tumor in the pelvis was discovered. She was admitted to our hospital for further examination of the pelvic tumor. She underwent laparotomy and was diagnosed with stage IIIA1 dysgerminoma (pT3apN0pM0) of the left ovary. Young female patients without the Y chromosome who are treated for primary amenorrhea may also develop malignant germ cell tumors; therefore, gynecologists should provide hormone replacement therapy and periodic pelvic evaluation.

A three-component model of the spinal nerve ramification: Bringing together the human gross anatomy and modern Embryology.

Due to its long history, the study of human gross anatomy has not adequately incorporated modern embryological findings; consequently, the current understanding has often been incompatible with recent discoveries from molecular studies. Notably, the traditional epaxial and hypaxial muscle distinction, and their corresponding innervation by the dorsal and ventral rami of the spinal nerve, do not correspond to the primaxial and abaxial muscle distinction, defined by the mesodermal lineages of target tissues. To resolve the disagreement between adult anatomy and embryology, we here propose a novel hypothetical model of spinal nerve ramification. Our model is based on the previously unknown developmental process of the intercostal nerves. Observations of these nerves in the mouse embryos revealed that the intercostal nerves initially had superficial and deep ventral branches, which is contrary to the general perception of a single ventral branch. The initial dual innervation pattern later changes into an adult-like single branch pattern following the retraction of the superficial branch. The modified intercostal nerves consist of the canonical ventral branches and novel branches that run on the muscular surface of the thorax, which sprout from the lateral cutaneous branches. We formulated the embryonic branching pattern into the hypothetical ramification model of the human spinal nerve so that the branching pattern is compatible with the developmental context of the target muscles. In our model, every spinal nerve consists of three components: (1) segmental branches that innervate the primaxial muscles, including the dorsal rami, and short branches and long superficial anterior branches from the ventral rami; (2) plexus-forming intramural branches, the serial homolog of the canonical intercostal nerves, which innervate the abaxial portion of the body wall; and (3) plexus-forming extramural branches, the series of novel branches located outside of the body wall, which innervate the girdle and limb muscles. The selective elaboration or deletion of each component successfully explains the reasoning for the standard morphology and variability of the spinal nerve. Therefore, our model brings a novel understanding of spinal nerve development and valuable information for basic and clinical sciences regarding the diverse branching patterns of the spinal nerve.

Epidemiology of human papillomavirus genotypes in pregnant Japanese women.

To investigate the pre-vaccination epidemiology of genital human papillomavirus (HPV) infections and genotypes in pregnant Japanese women, we performed Pap smear tests and HPV genotype testing in patients attending Nagasaki University Hospital and collaborating hospitals from August 2007 to July 2010. Serial uterine cervical specimens were obtained from 151 pregnant women. The HPV test was positive on the first visit in 54 women (35.8%; 54/151, average age 30). A total of 49 women (32.5%; 49/151) were infected by at least one high-risk HPV and 5 women were infected by only low-risk HPV. The three most prevalent high-risk HPV genotypes were HPV 52 (31.5%; 17/54), HPV 16 (29.6%; 16/51) and HPV 31 (13.0%; 7/51). The HPV infection pattern (negative, single infection and multiple infection) differed significantly according to the pregnancy trimester (χ(2)-test; P<0.01(Pearson)). Among HPV-infected pregnant Japanese women, HPV52 was the most common genotype. The second most common genotype was HPV16, and these two genotypes accounted for ∼60% of HPV-positive pregnant women. Infection with multiple HPV genotypes was observed more frequently in the first trimester of pregnancy and the pattern of infection changed significantly depending on pregnancy stage.

Clinical application of fetal sex determination using cell-free fetal DNA in pregnant carriers of X-linked genetic disorders.

As the first step in prenatal diagnosis of X-linked genetic disorders, chorionic villus sampling (CVS) for fetal sex determination is generally performed at 11-13 weeks of gestation. However, as the procedure-related miscarriage rate of CVS is 0.5-1.0%, non-invasive methods such as PCR of cell-free fetal DNA (cff-DNA) in maternal plasma are preferable. Here, we determined fetal sex at 9-12 weeks of gestation using PCR of cff-DNA in three pregnant carriers of Duchenne muscular dystrophy. The fetal sex was accurately determined in all three cases, as confirmed by ultrasound and amniocentesis at 16 weeks (for the two female fetuses) and CVS at 12 weeks (for the one male fetus). This procedure could avoid unnecessary CVS in female fetuses.

Pre-vaccination epidemiology of human papillomavirus infections in Japanese women with abnormal cytology.

AIM: To investigate the pre-vaccination epidemiology of genital human papillomavirus (HPV) infections and genotypes in women with abnormal cytology in Nagasaki, Japan. MATERIAL AND METHODS: We performed Pap smear tests, biopsies and HPV genotype testing in Nagasaki Prefecture from August 2007 through November 2009. RESULTS: During the study period, serial samples of uterine cervical specimens were obtained from 539 subjects with abnormal cytology and/or squamous intraepithelial lesions (SIL) confirmed previously, or with clinically suspected invasive cervical cancer. In 119 HPV-positive subjects with low-grade SIL, the three most prevalent high-risk HPV genotypes were HPV52 (21.8%; 26/119), HPV16 (20.2%; 24/119) and HPV56 (17.6%; 21/119). In 199 women, 127 HPV-positive subjects with high-grade SIL and 67 HPV-positive subjects with squamous cell carcinoma (SCC), the three most prevalent high-risk HPV genotypes were HPV16 (44.3%; 86/194), HPV52 (20.6%; 40/194) and HPV58 (16.0%; 31/194). CONCLUSION: Compared with the distribution of high-risk HPV genotypes in other countries, HPV52 was a more common genotype in Nagasaki. With disease progression to SCC, the distribution of high-risk HPV56 belonging to the A6 HPV family decreased, while HPV16 and HPV52 belonging to the A9 HPV family persisted. Our data provide an important resource to address the case for vaccination against HPV genotypes other than HPV16 and HPV18 in Japan.

Human papillomavirus DNA in plasma of patients with HPV16 DNA-positive uterine cervical cancer.

OBJECTIVES: The squamous cell carcinoma antigen is considered the most accurate serologic tumor marker for uterine cervical carcinoma. However, serum squamous cell carcinoma antigen levels were found to correlate significantly with clinical severity of atopic dermatitis and chronic renal failure. The present study was conducted in patients with human papillomavirus 16 DNA-positive uterine cervical cancer to determine the plasma level of human papillomavirus 16 DNA and the diagnostic values of plasma human papillomavirus DNA in these patients. METHODS: Forty-three human papillomavirus 16-positive patients with cervical intraepithelial neoplasia or uterine cervical squamous cell carcinoma were recruited in this study. The diagnosis was cervical cancer in 20 patients, high-grade squamous intraepithelial lesions in 21, low-grade squamous intraepithelial lesions in 1 and negative for intraepithelial lesion or malignancy in 3 patients. Before any treatment, blood samples were collected from all patients. For analysis of human papillomavirus DNA in plasma of patients with cervical cancer, quantitative polymerase chain reaction fluorescent assay for human papillomavirus 16 was performed using human papillomavirus 16 primers and SYBR Green dye using the LightCycler 480 SW1.5 apparatus. RESULTS: Plasma human papillomavirus 16 DNA was detected in only 30.0% of the patients with human papillomavirus 16-positive cervical cancer and in none of normal controls. The copy number of plasma human papillomavirus 16 DNA was higher in patients with invasive cancer than in those with cervical intraepithelial neoplasia (CIN3), micro-invasive cancer and in normal individuals. CONCLUSIONS: These results indicated that the plasma human papillomavirus DNA level could be potentially used as a marker of low-invasive cervical cancer tumors in patients with normal squamous cell carcinoma antigen levels before treatment.

The possibility of microarray-based analysis using cell-free placental mRNA in maternal plasma.

OBJECTIVE: The purpose of this study is to investigate a possibility of overall assessment of cell-free (CF) placental mRNAs in maternal plasma. METHODS: First, placenta-predominantly expressed transcripts were selected by the analysis of GeneChip using three sets of placental tissues and corresponding maternal blood cells. Subsequently, a custom cDNA array panel of placenta-predominantly expressed transcripts was designed and used to compare the RNA profiles of maternal plasma collected from 12 preeclamptic and 12 uncomplicated pregnancies. Scatter plots for the signal intensities of the comparative cDNA hybridization revealed either unchanged or aberrant patterns. RESULTS: We selected top 50 placenta-predominantly expressed transcripts that were > 2500 times higher in placental tissues than in corresponding whole blood samples. A custom cDNA array analysis detected the aberrant pattern in five preeclamptic women with severe hypertension but not in seven preeclamptic women with mild hypertension (P < 0.05, Fisher's direct method). The aberrant pattern of above RNA transcripts in maternal plasma was validated by quantitative real-time reverse transcription-polymerase chain reaction. The mean (range) value of coefficient of variations in this custom array quantification was 9.4% (3.0-16.2%). CONCLUSION: Our custom cDNA array is expected to be useful for overall assessment of CF placental mRNAs in maternal plasma in a single experiment.

Development of enzyme-linked immunosorbent assay for detection of antibody against Caprine arthritis encephalitis virus using recombinant protein of the precursor of the major core protein, p55gag.

An indirect enzyme-linked immunosorbent assay (ELISA) by using recombinant Caprine arthritis encephalitis virus (CAEV) p55gag antigen (rELISA), an indirect ELISA by using whole CAEV (wELISA), and Western blot analysis by using the recombinant p55gag antigen (rWB) were developed for detection of CAEV-specific antibodies in goats. Seven hundred and forty-five sera from goats were tested by rELISA, wELISA, rWB, and agar gel immunodiffusion test (AGID), and the results were compared with those of WB analysis by using the whole CAEV antigen (wWB). The AGID test and rWB had similar sensitivities of 93.3% (95% confidence interval [CI]) and 93% (95% CI), respectively, and similar specificities of 96.0% (95% CI) and 96.3% (95% CI), respectively, compared with wWB. The wELISA had substantially lower sensitivity (80.4%) and specificity (78.0%) compared with wWB, and rELISA had the lowest sensitivity (78.2%) and specificity (61.1%) compared with wWB. The lack of adequate sensitivity and specificity for rELISA and wELISA suggests that these assays need considerable modification. However, the results for rWB show that this assay has excellent agreement with wWB and that it can be used as a confirmatory test for the presence of anti-CAEV antibodies.

Placental multiple chorionic cysts in maternal scleroderma.

We present a case of a pregnant woman with scleroderma (Ssc) whose placenta showed multiple chorionic cysts and severe fibrotic changes and large infarcted areas at the maternal side. Fetal growth was appropriate for gestational age and amniotic fluid volume was normal, but fetal tachycardia, loss of variability, and late deceleration were detected by non-stress test at 29 weeks of gestation. She was diagnosed as having non-reassuring fetal status and delivered a female baby who weighed 1092 g (Apgar score 6/9) by Caesarean section. Placental surface cysts are rare findings and their effect on pregnancy is poorly understood, but an association with intrauterine growth restriction (IUGR) has been reported. This is the first report of a pregnant woman with scleroderma showing multiple placental cysts.

Expression pattern of LRR and Ig domain-containing protein (LRRIG protein) in the early mouse embryo.

The combination of leucine-rich repeat (LRR) and immunoglobulin-like (Ig) domains is found in the domain architecture of the Trk neurotrophin receptor protein. Recently dozens of such proteins simultaneously carrying LRR and Ig domains as the Trk receptors have been identified. Given the significant biological roles of Trk and such newly identified proteins, we have searched the public database for human proteins with LRR and Ig domains (collectively termed the leucine-rich repeat and Ig domain-containing protein, LRRIG protein, in this study), and have analyzed the mRNA expression pattern of mouse orthologs of obtained human LRRIG proteins at embryonic day 10. The list of the LRRIG proteins includes 36 human proteins: four LINGO, three NGL, five SALM, three NLRR, three Pal, two ISLR, three LRIG, two GPR, two Adlican, two Peroxidasin-like proteins, three Trk neurotrophin receptors, a yet unnamed protein AAI11068, and three AMIGO. Some molecules (LINGO2, LINGO4, NGL1, SALM1, SALM5, and TrkB) were expressed exclusively in neuronal tissues, whereas others (ISLR1, GPR124, and Adlican2) exhibited non-neuronal expression profiles. However, the majority of LRRIG protein family exhibited broad mRNA tissue-expression profiles.

Molecular properties of the putative autolysin Atl(WM) encoded by Staphylococcus warneri M: mutational and biochemical analyses of the amidase and glucosaminidase domains.

The putative autolysin Atl(WM) of Staphylococcus warneri M is a modular protein exhibiting two enzyme activities, an N-terminal side amidase (ami(atlwm)-R1-R2) and a C-terminal side glucosaminidase (R3-glu(atlwm)). Zymographic analysis of the protein overproduced in Escherichia coli showed that both enzymes were active toward 17 Gram-positive bacteria, including staphylococci, lactobacilli, lactococci, enterococci, and micrococci. The purified enzyme core ami(atlwm) (or glu(atlwm)) had the pH and temperature optima of about 7.0 (5.5) and 41 (50) degrees C, respectively. ami(atlwm) was inactivated by EDTA, and was stimulated by such salts as CoCl(2), MnCl(2), CaCl(2), or ZnCl(2). Six mutations within ami(atlwm), (H362A, E421A, H467A, H479, D481A, and Y491D) drastically reduced cell-lytic activity. Comparative analysis with other related amidases suggested that the three residues H362, H467, and D481 likely act as ligands (and/or active sites). The lytic activity of glu(atlwm) markedly declined in four mutants (E1238A, E1238Q, T1239A, and Y1332A). For determination of the putative cell-recognition regions, four domains (R1-R2, R1, R2, and R3) were purified; all the proteins substantially bound to S. warneri M cells from exponential to stationary growth phases, and R1-R2 aggregated the cells. Protein sequencing and immunoblot analysis suggested that the extacellular Atl(WM) might be primarily processed at two specific sites (one between pro and ami(atlwm), and the other between R2 and R3) to yield the mature amidase and glucosaminidase.