Author correction: the solute carrier SLC9C1 is a Na(+)/H(+)-exchanger gated by an S4-type voltage-sensor and cyclic-nucleotide binding

© The Author(s), 2020. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Windler, F., Bönigk, W., Körschen, H. G., Grahn, E., Strünker, T., Seifert, R., & Kaupp, U. B. Author correction: the solute carrier SLC9C1 is a Na(+)/H(+)-exchanger gated by an S4-type voltage-sensor and cyclic-nucleotide binding. Nature Communications, 11(1),(2020): 4210, doi:10.1038/s41467-020-18023-5

Funktionelle Charakterisierung eines spermienspezifischen Na⁺/H⁺-Austauschers

Sekundär aktive Austauscher sind membranständige Proteine, die den elektrochemischen Gradienten eines Ions nutzen, um den Transport anderer Ionen oder Substrate über die Zellmembran zu ermöglichen. Natrium/Protonen (Na+/H+) Austauscher der Solute Carrier 9 (SLC9) Familie spielen dabei eine Rolle in einer Vielzahl von physiologischen Prozessen. Ein bis heute rätselhaftes Mitglied der SLC9 Familie ist der spermienspezifische Austauscher SLC9C1. Es ist bis heute nicht bekannt, ob SLC9C1 ein funktionstüchtiger Na+/H+ Austauscher ist. SLC9C1 besitzt eine einzigartige dreiteilige Struktur; eine Austauscher (NHE) Domäne, eine Spannungssensordomäne (VSD) sowie eine Bindestelle für zyklische Nukleotide (CNBD). VSDs und CNBDs sind funktionelle Proteindomänen, die man bereits in anderen Proteinen gefunden hat, VSDs überwiegend in Ionenkanälen, aber auch in einer spannungsabhängigen Phosphatase, CNBDs in Proteinkinasen, Transkriptionsfaktoren, und auch in Ionenkanälen. SLC9C1 ist das erste Mitglied der SLC Familie, in dem solche Proteindomänen gefunden wurden. Im Rahmen meiner Arbeit habe ich diesen phylogenetischen Hybrid untersucht. Als Modellprotein diente dazu SLC9C1 des Seeigels Strongylocentrotus purpuratus (SpSLC9C1). Durch den Einsatz pH sensitiver Fluoreszenzfarbstoffe in der Elektrophysiologie konnte ich zeigen, dass SpSLC9C1 ein funktionstüchtiger Na+/H+ Austauscher ist. Hyperpolarisierende Spannungssprünge aktivieren sowohl Gating Ströme der VSD als auch den sekundär aktiven Na+/H+ Austausch. Ich konnte die Spannungsabhängigkeit der Austauschaktivität bestimmen, sowie zeigen, dass cAMP, aber nicht cGMP, die Spannungsabhängigkeit der Gating Ströme und der Austauschaktivität moduliert. Mutationen in der NHE Domäne, der CNBD und der VSD verhinderten jeweils den Na+/H+ Austausch bzw. die Modulation durch cAMP oder beeinflussten die Gating Ströme. Ein Antikörper bestätigte die Expression von SLC9C1 in S. purpuratus Spermien. Mithilfe von spannungs , pH und Na+ sensitiven Fluoreszenzfarbstoffen in S. purpuratus Spermien konnte ich zeigen, dass SpSLC9C1 auch in Spermien durch Hyperpolarisation aktiviert wird. Der Na+/H+ Austausch alkalisiert das Flagellum nach der Lockstoffstimulation und ermöglicht dem Spermium die chemotaktische Navigation.The solute carrier SLC9C1 is a Na+/H+-exchanger gated by an S4-type voltage-sensor and cyclic-nucleotide binding Secondary active exchangers are membrane bound proteins that use the electrochemical gradient of ions to transport other ions or substrates across cell membranes. Within these proteins, sodium/proton (Na+/H+) exchangers of the Solute Carrier 9 (SLC9) family play a crucial role in a number of physiological processes. A so far enigmatic member of this family is the sperm specific exchanger SLC9C1. No evidence exists up to today that SLC9C1 is a functional Na+/H+ exchanger. SLC9C1 features a unique tripartite structure; an exchanger (NHE) domain, a voltage sensing domain (VSD) as well as a cyclic nucleotide binding domain (CNBD). VSDs and CNBDs are protein domains that are found in a variety of proteins, VSDs mainly in ion channels but also in a voltage sensitive phosphatase, CNBDs in protein kinases, transcription factors and also ion channels. SLC9C1 is the first member of the SLC9 family in which these protein domains was found. During my thesis, I investigated this phylogenetic hybrid. As a model protein, I chose SLC9C1 from the sea urchin Strongylocentrotus purpuratus (SpSLC9C1). By using a pH sensitive fluorescence dye within an electrophysiological setup, I could show that SpSLC9C1 is indeed a functional Na+/H+ exchanger. Hyperpolarizing voltage steps evoked both gating currents and secondary active Na+/H+ exchange. I could determine the voltage dependence of exchanger activation. Furthermore, I looked into the action of cyclic nucleotides such as cGMP and cAMP on SpSLC9C1. Cyclic AMP, but not cGMP affected the voltage dependence of both gating currents and exchange activity. Mutations in the NHE domain, CNBD and VSD abolished Na+/H+ exchange, cAMP modulation or affected voltage gating, respectively. An antibody was raised to confirm SLC9C1 expression in S. purpuratus sperm. By using voltage , pH and Na+ sensitive fluorescent dyes together with a rapid mixing device, I could show that SpSLC9C1 in sperm activates during hyperpolarization. The Na+/H+ exchange alkalizes the flagellum upon stimulation with the chemoattractant and allows chemotactic navigation

Structural and biophysical characterization of the type VII collagen vWFA2 subdomain leads to identification of two binding sites

Type VII collagen is an extracellular matrix protein, which is important for skin stability; however, detailed information at the molecular level is scarce. The second vWFA (von Willebrand factor type A) domain of type VII collagen mediates important interactions, and immunization of mice induces skin blistering in certain strains. To understand vWFA2 function and the pathophysiological mechanisms leading to skin blistering, we structurally characterized this domain by X-ray crystallography and NMR spectroscopy. Cell adhesion assays identified two new interactions: one with beta 1 integrin via its RGD motif and one with laminin-332. The latter interaction was confirmed by surface plasmon resonance with a K-D of about 1 mm. These data show that vWFA2 has additional functions in the extracellular matrix besides interacting with type I collagen

Perimenopause and Postmenopause - Diagnosis and Interventions. Guideline of the DGGG and OEGGG (S3-Level, AWMF Registry Number 015-062, September 2020).

Aim The aim of the interdisciplinary S3-guideline Perimenopause and Postmenopause - Diagnosis and Interventions is to provide help to physicians as they inform women about the physiological changes which occur at this stage of life and the treatment options. The guideline should serve as a basis for decisions taken during routine medical care. This short version lists the statements and recommendations given in the long version of the guideline together with the evidence levels, the level of recommendation, and the strength of consensus. Methods The statements and recommendations are largely based on methodologically high-quality publications. The literature was evaluated by experts and mandate holders using evidence-based medicine (EbM) criteria. The search for evidence was carried out by the Essen Research Institute for Medical Management (EsFoMed). To some extent, this guideline also draws on an evaluation of the evidence used in the NICE guideline on Menopause and the S3-guidelines of the AWMF and has adapted parts of these guidelines. Recommendations Recommendations are given for the following subjects: diagnosis and therapeutic interventions for perimenopausal and postmenopausal women, urogynecology, cardiovascular disease, osteoporosis, dementia, depression, mood swings, hormone therapy and cancer risk, as well as primary ovarian insufficiency