The mitogen-activated protein kinase (MAPK) pathway is the canonical signaling pathway for many receptor tyrosine kinases, such as the Epidermal Growth Factor Receptor. Downstream of the receptors, this pathway involves the activation of a kinase cascade that culminates in a transcriptional response and affects processes, such as cell migration and adhesion. In addition, the strength and duration of the upstream signal also influence the mode of the cellular response that is switched on. Thus, the same components can in principle coordinate opposite responses, such as proliferation and differentiation. In recent years, it has become evident that MAPK signaling is regulated and fine-tuned by proteins that can bind to several MAPK signaling proteins simultaneously and, thereby, affect their function. These so-called MAPK scaffolding proteins are, thus, important coordinators of the signaling response in cells. In this review, we summarize the recent advances in the research on MAPK/extracellular signal-regulated kinase (ERK) pathway scaffolders. We will not only review the well-known members of the family, such as kinase suppressor of Ras (KSR), but also put a special focus on the function of the recently identified or less studied scaffolders, such as fibroblast growth factor receptor substrate 2, flotillin-1 and mitogen-activated protein kinase organizer 1.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated sequence 9 (CRISPR/Cas9) system is widely used for genome editing purposes as it facilitates an efficient knockout of a specific gene in, e.g. cultured cells. Targeted double-strand breaks are introduced to the target sequence of the guide RNAs, which activates the cellular DNA repair mechanism for non-homologous-end-joining, resulting in unprecise repair and introduction of small deletions or insertions. Due to this, sequence alterations in the coding region of the target gene frequently cause frame-shift mutations, facilitating degradation of the mRNA. We here show that such CRISPR/Cas9-mediated alterations in the target exon may also result in altered splicing of the respective pre-mRNA, most likely due to mutations of splice-regulatory sequences. Using the human FLOT-1 gene as an example, we demonstrate that such altered splicing products also give rise to aberrant protein products. These may potentially function as dominant-negative proteins and thus interfere with the interpretation of the data generated with these cell lines. Since most researchers only control the consequences of CRISPR knockout at genomic and protein level, our data should encourage to also check the alterations at the mRNA level.

Flotillins are highly conserved proteins that localize into specific cholesterol rich microdomains in cellular membranes. They have been shown to be associated with, for example, various signaling pathways, cell adhesion, membrane trafficking and axonal growth. Recent findings have revealed that flotillins are frequently overexpressed in various types of human cancers. We here review the suggested functions of flotillins during receptor tyrosine kinase signaling and in cancer. Although flotillins have been implicated as putative cancer therapy targets, we here show that great caution is required since flotillin ablation may result in effects that increase instead of decrease the activity of specific signaling pathways. On the other hand, as flotillin overexpression appears to be related with metastasis formation in certain cancers, we also discuss the implications of these findings for future therapy aspects.

Succinic semialdehyde dehydrogenase deficiency (SSADH-D) is a genetic disorder that results from the aberrant metabolism of the neurotransmitter γ-amino butyric acid (GABA). The disease is caused by impaired activity of the mitochondrial enzyme succinic semialdehyde dehydrogenase. SSADH-D manifests as varying degrees of mental retardation, autism, ataxia, and epileptic seizures, but the clinical picture is highly heterogeneous. So far, there is no approved curative therapy for this disease. In this review, we briefly summarize the molecular genetics of SSADH-D, the past and ongoing clinical trials, and the emerging features of the molecular pathogenesis, including redox imbalance and mitochondrial dysfunction. The main aim of this review is to discuss the potential of further therapy approaches that have so far not been tested in SSADH-D, such as pharmacological chaperones, read-through drugs, and gene therapy. Special attention will also be paid to elucidating the role of patient advocacy organizations in facilitating research and in the communication between researchers and patients.

Acetylcholine is an important neurotransmitter whose effects are mediated by two classes of receptors. The nicotinic acetylcholine receptors are ion channels, whereas the muscarinic receptors belong to the large family of G protein coupled seven transmembrane helix receptors. Beyond its function in neuronal systems, it has become evident that acetylcholine also plays an important role in non-neuronal cells such as epithelial and immune cells. Furthermore, many cell types in the periphery are capable of synthesizing acetylcholine and express at least some of the receptors. In this review, we summarize the non-neuronal functions of the muscarinic acetylcholine receptors, especially those of the M₂ muscarinic receptor in epithelial cells. We will review the mechanisms of signaling by the M₂ receptor but also the cellular trafficking and ARF6 mediated endocytosis of this receptor, which play an important role in the regulation of signaling events. In addition, we provide an overview of the M₂ receptor in human pathological conditions such as autoimmune diseases and cancer.

Pemphigus Vulgaris is an autoimmune disease that results in blister formation in the epidermis and in mucosal tissues due to antibodies recognizing desmosomal cadherins, mainly desmoglein-3 and -1. Studies on the molecular mechanisms of Pemphigus have mainly been carried out using the spontaneously immortalized human keratinocyte cell line HaCaT or in primary keratinocytes. However, both cell systems have suboptimal features, with HaCaT cells exhibiting a large number of chromosomal aberrations and mutated p53 tumor suppressor, whereas primary keratinocytes are short-lived, heterogeneous and not susceptible to genetic modifications due to their restricted life-span. We have here tested the suitability of the commercially available human keratinocyte cell line hTert/KER-CT as a model system for research on epidermal cell adhesion and Pemphigus pathomechanisms. We here show that hTert cells exhibit a calcium dependent expression of desmosomal cadherins and are well suitable for typical assays used for studies on Pemphigus, such as sequential detergent extraction and Dispase-based dissociation assay. Treatment with Pemphigus auto-antibodies results in loss of monolayer integrity and altered localization of desmoglein-3, as well as loss of colocalization with flotillin-2. Our findings demonstrate that hTert cells are well suitable for studies on epidermal cell adhesion and Pemphigus pathomechanisms.