Using for High-Yield Production of Challenging Expression Targets and for Protein Perdeuteration.

Production of soluble proteins is essential for structure/function studies; however, this usually requires milligram amounts of protein, which can be difficult to obtain with traditional expression systems. Recently, the Gram-negative bacterium emerged as a novel and alternative host platform for production of proteins in high yields. Here, we used a commercial strain derived from (Vmax X2) to produce soluble bacterial and fungal proteins in milligram scale, which we struggled to achieve in .

Using for high-yield production of challenging expression targets and for protein deuteration.

Production of soluble proteins is essential for structure/function studies, however, this usually requires milligram amounts of protein, which can be difficult to obtain with traditional expression systems. Recently, the Gram-negative bacterium appeared as a novel and alternative host platform for production of proteins in high yields. Here, we used a commercial strain derived from (Vmax X2) to produce soluble bacterial and fungal proteins in milligram scale, which we struggled to achieve in .

CYSRT1: An Antimicrobial Epidermal Protein that Can Interact with Late Cornified Envelope Proteins.

Late cornified envelope (LCE) proteins are small cationic epidermal proteins with antimicrobial properties, and the combined deletion of LCE3B and LCE3C genes is a risk factor for psoriasis that affects skin microbiome composition. In a yeast two-hybrid screen, we identified CYSRT1 as an interacting partner of members of all LCE groups except LCE6. These interactions were confirmed in a mammalian cell system by coimmunoprecipitation.

Cathepsin B as a potential cystatin M/E target in the mouse hair follicle.

Deficiency of the cysteine protease inhibitor cystatin M/E (Cst6) in mice leads to disturbed epidermal cornification, impaired barrier function, and neonatal lethality. We report the rescue of the lethal skin phenotype of (Cst6-deficient; ) mice by transgenic, epidermis-specific, reexpression of Cst6 under control of the human involucrin (INV) promoter. Rescued Tg(INV-) mice survive the neonatal phase, but display severe eye pathology and alopecia after 4 mo.

Implementing targeted expectant management in fertility care using prognostic modelling: a cluster randomized trial with a multifaceted strategy.

Study Question: What is the effectiveness of a multifaceted implementation strategy compared to usual care on improving the adherence to guideline recommendations on expectant management for couples with unexplained infertility?

Summary Answer: The multifaceted implementation strategy did not significantly increase adherence to guideline recommendations on expectant management compared to care as usual.

What Is Known Already: Intrauterine insemination (IUI) with or without ovarian hyperstimulation has no beneficial effect compared to no treatment for 6 months after the fertility work-up for couples with unexplained infertility and a good prognosis of natural conception. Therefore, various professionals and policy makers have advocated the use of prognostic profiles and expectant management in guideline recommendations.

Cost-effectiveness of 'immediate IVF' versus 'delayed IVF': a prospective study.

Study Question: How does the cost-effectiveness (CE) of immediate IVF compared with postponing IVF for 1 year, depend on prognostic characteristics of the couple?

Summary Answer: The CE ratio, i.e. the incremental costs of immediate versus delayed IVF per extra live birth, is the highest (range of €15 000 to >€60 000) for couples with unexplained infertility and for them depends strongly on female age and the duration of infertility, whilst being lowest for endometriosis (range 8000-23 000) and, for such patients, only slightly dependent on female age and duration of infertility.

["STeP" ("Students Teaching Patients"): Medical Students Following up on Informed Consent Discussions. Results of a Methodological Change and Further Considerations].

Literature shows an increasing number of reports on the incompleteness of informed consent discussions held by residents. Residents feel insecure and not adequately prepared for this task. This project aimed to integrate supervised informed consent discussions into the medical curriculum, working with "real" patients instead of other students or actors.

Tailored expectant management in couples with unexplained infertility does not influence their experiences with the quality of fertility care.

Study Question: Do couples who were eligible for tailored expectant management (TEM) and did not start treatment within 6 months after the fertility work-up, have different experiences with the quality of care than couples that were also eligible for TEM but started treatment right after the fertility work-up?

Summary Answer: Tailored expectant management of at least 6 months in couples with unexplained infertility is not associated with the experiences with quality of care or trust in their physician.

What Is Known Already: In couples with unexplained infertility and a good prognosis of natural conception within 1 year, expectant management for 6-12 months does not compromise ongoing birth rates and is equally as effective as starting medically assisted reproduction immediately. Therefore, TEM is recommended by various international clinical guidelines.

Late cornified envelope (LCE) proteins: distinct expression patterns of LCE2 and LCE3 members suggest nonredundant roles in human epidermis and other epithelia.

Background: Deletion of the late cornified envelope (LCE) proteins LCE3B and LCE3C is a strong and widely replicated psoriasis risk factor. It is amenable to biological analysis because it precludes the expression of two epidermis-specific proteins, rather than being a single-nucleotide polymorphism of uncertain significance. The biology of the 18-member LCE family of highly homologous proteins has remained largely unexplored so far.

[Overtreatment in couples with unexplained subfertility].

Objective: To assess adherence to expectant management of 6-12 months in couples with unexplained subfertility, as recommended by the Dutch Networkguideline Subfertility.

Design: A retrospective cohort study in 25 clinics.

Method: Couples were eligible to participate if they were diagnosed with unexplained subfertility and had a good prognosis of natural conception within one year (>30%), for these couples the network guideline recommends an expectant management.

Overtreatment in couples with unexplained infertility.

Study Question: What is the percentage of overtreatment, i.e. fertility treatment started too early, in couples with unexplained infertility who were eligible for tailored expectant management?

Summary Answer: Overtreatment occurred in 36% of couples with unexplained infertility who were eligible for an expectant management of at least 6 months.

Improving the implementation of tailored expectant management in subfertile couples: protocol for a cluster randomized trial.

Background: Prognostic models in reproductive medicine can help to identify subfertile couples who would benefit from fertility treatment. Expectant management in couples with a good chance of natural conception, i.e.

The mitotic spindle protein SPAG5/Astrin connects to the Usher protein network postmitotically.

Background: Mutations in the gene for Usher syndrome 2A (USH2A) are causative for non-syndromic retinitis pigmentosa and Usher syndrome, a condition that is the most common cause of combined deaf-blindness. To gain insight into the molecular pathology underlying USH2A-associated retinal degeneration, we aimed to identify interacting proteins of USH2A isoform B (USH2AisoB) in the retina.

Results: We identified the centrosomal and microtubule-associated protein sperm-associated antigen (SPAG)5 in the retina.

Cystatin M/E knockdown by lentiviral delivery of shRNA impairs epidermal morphogenesis of human skin equivalents.

The protease inhibitor cystatin M/E (CST6) regulates a biochemical pathway involved in stratum corneum homeostasis, and its deficiency in mice causes ichthyosis and neonatal lethality. Cystatin M/E deficiency has not been described in humans so far, and we did not detect disease-causing mutations in the CST6 gene in a large number of patients with autosomal recessive congenital ichthyosis, who were negative for mutations in known ichthyosis-associated genes. To investigate the phenotype of CST6 deficiency in human epidermis, we used lentiviral delivery of short hairpin RNAs that target CST6 in a 3D reconstructed skin model.

CLRN1 mutations cause nonsyndromic retinitis pigmentosa.

Objective: To describe the mutations in the CLRN1 gene in patients from 2 consanguineous Pakistani families diagnosed with autosomal recessive retinitis pigmentosa (arRP).

Design: Case-series study.

Participants: Affected and unaffected individuals of 2 consanguineous Pakistani families and 90 unaffected controls from the same population.

Sequence variants of the DFNB31 gene among Usher syndrome patients of diverse origin.

Purpose: It has been demonstrated that mutations in deafness, autosomal recessive 31 (DFNB31), the gene encoding whirlin, is responsible for nonsyndromic hearing loss (NSHL; DFNB31) and Usher syndrome type II (USH2D). We screened DFNB31 in a large cohort of patients with different clinical subtypes of Usher syndrome (USH) to determine the prevalence of DFNB31 mutations among USH patients.

Methods: DFNB31 was screened in 149 USH2, 29 USH1, six atypical USH, and 11 unclassified USH patients from diverse ethnic backgrounds.

Association of whirlin with Cav1.3 (alpha1D) channels in photoreceptors, defining a novel member of the usher protein network.

Purpose: Usher syndrome is the most common form of hereditary deaf-blindness. It is both clinically and genetically heterogeneous. The USH2D protein whirlin interacts via its PDZ domains with other Usher-associated proteins containing a C-terminal type I PDZ-binding motif.

Usher syndrome and Leber congenital amaurosis are molecularly linked via a novel isoform of the centrosomal ninein-like protein.

Usher syndrome (USH) and Leber congenital amaurosis (LCA) are autosomal recessive disorders resulting in syndromic and non-syndromic forms of blindness. In order to gain insight into the pathogenic mechanisms underlying retinal degeneration, we searched for interacting proteins of USH2A isoform B (USH2A(isoB)) and the LCA5-encoded protein lebercilin. We identified a novel isoform of the centrosomal ninein-like protein, hereby named Nlp isoform B (Nlp(isoB)), as a common interactor.

A novel Usher protein network at the periciliary reloading point between molecular transport machineries in vertebrate photoreceptor cells.

The human Usher syndrome (USH) is the most frequent cause of combined deaf-blindness. USH is genetically heterogeneous with at least 12 chromosomal loci assigned to three clinical types, USH1-3. Although these USH types exhibit similar phenotypes in human, the corresponding gene products belong to very different protein classes and families.

MPP1 links the Usher protein network and the Crumbs protein complex in the retina.

The highly ordered distribution of neurons is an essential feature of a functional mammalian retina. Disruptions in the apico-basal polarity complexes at the outer limiting membrane (OLM) of the retina are associated with retinal patterning defects in vertebrates. We have analyzed the binding repertoire of MPP5/Pals1, a key member of the apico-basal Crumbs polarity complex, that has functionally conserved counterparts in zebrafish (nagie oko) and Drosophila (Stardust).

Mutations in LCA5, encoding the ciliary protein lebercilin, cause Leber congenital amaurosis.

Leber congenital amaurosis (LCA) causes blindness or severe visual impairment at or within a few months of birth. Here we show, using homozygosity mapping, that the LCA5 gene on chromosome 6q14, which encodes the previously unknown ciliary protein lebercilin, is associated with this disease. We detected homozygous nonsense and frameshift mutations in LCA5 in five families affected with LCA.

Mutations in the lipoma HMGIC fusion partner-like 5 (LHFPL5) gene cause autosomal recessive nonsyndromic hearing loss.

In two large Turkish consanguineous families, a locus for autosomal recessive nonsyndromic hearing loss (ARNSHL) was mapped to chromosome 6p21.3 by genome-wide linkage analysis in an interval overlapping with the loci DFNB53 (COL11A2), DFNB66, and DFNB67. Fine mapping excluded DFNB53 and subsequently homozygous mutations were identified in the lipoma HMGIC fusion partner-like 5 (LHFPL5) gene, also named tetraspan membrane protein of hair cell stereocilia (TMHS) gene, which was recently shown to be mutated in the "hurry scurry" mouse and in two DFNB67-linked families from Pakistan.

The DFNB31 gene product whirlin connects to the Usher protein network in the cochlea and retina by direct association with USH2A and VLGR1.

Mutations in the DFNB31 gene encoding the PDZ scaffold protein whirlin are causative for hearing loss in man and mouse. Whirlin is known to be essential for the elongation process of the stereocilia of sensory hair cells in the inner ear, though its complete spatial and temporal expression patterns remained elusive. Here, we demonstrate that, in embryonic development, the gene is not only expressed in the inner ear, but also in the developing brain and the retina.