The evolution of proteins is primarily driven by the combinatorial assembly of a limited set of pre-existing modules known as protein domains. This modular architecture not only supports the diversity of natural proteins but also provides a robust strategy for protein engineering, enabling the design of artificial proteins with enhanced or novel functions for various industrial applications. Among these functions, oligomerization plays a crucial role in enhancing protein activity, such as by increasing the binding capacity of antibodies. To investigate the potential of engineering oligomerization, we examined the transferability of the sequence domain encoded by exon 5 (Ex5), which was originally responsible for the oligomerization of ameloblastin (AMBN). We designed a two-domain protein composed of Ex5 in combination with a monomeric, globular, and highly stable protein, specifically calmodulin (CaM). CaM represents the opposite protein character to AMBN, which is highly disordered and has a dynamic character. This engineered protein, termed eCaM, successfully acquired an oligomeric function, inducing self-assembly under specific conditions. Biochemical and biophysical analyses revealed that the oligomerization of eCaM is both concentration- and time-dependent, with the process being reversible upon dilution. Furthermore, mutating a key oligomerization residue within Ex5 abolished the self-assembly of eCaM, confirming the essential role of the Ex5 motif in driving oligomerization. Our findings demonstrate that the oligomerization properties encoded by Ex5 can be effectively transferred to a new protein context, though the positioning of Ex5 within the protein structure is critical. This work highlights the potential of enhancing monomeric proteins with oligomeric functions, paving the way for industrial applications and the development of proteins with tailored properties.

• Publikační typ
• časopisecké články MeSH

The role of glia in amyotrophic lateral sclerosis (ALS) is undeniable. Their disease-related activity has been extensively studied in the spinal cord, but only partly in the brain. We present herein a comprehensive study of glia in the cortex of SOD1(G93A) mice-a widely used model of ALS. Using single-cell RNA sequencing (scRNA-seq) and immunohistochemistry, we inspected astrocytes, microglia, and oligodendrocytes, in four stages of the disease, respecting the factor of sex. We report minimal changes of glia throughout the disease progression and regardless of sex. Pseudobulk and single-cell analyses revealed subtle disease-related transcriptional alterations at the end-stage in microglia and oligodendrocytes, which were supported by immunohistochemistry. Therefore, our data support the hypothesis that the SOD1(G93A) mouse cortex does not recapitulate the disease in patients, and we recommend the use of a different model for future studies of the cortical ALS pathology.

• MeSH
• amyotrofická laterální skleróza * genetika   patologie   MeSH
• mícha patologie   MeSH
• modely nemocí na zvířatech MeSH
• motorické neurony patologie   MeSH
• myši transgenní MeSH
• myši MeSH
• neuroglie * patologie   MeSH
• superoxiddismutasa 1 * genetika   MeSH
• superoxiddismutasa genetika   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• superoxiddismutasa 1 * MeSH
• superoxiddismutasa MeSH

Engineering artificial skin constructs is an ongoing challenge. An ideal material for hosting skin cells is still to be discovered. A promising candidate is low-cost cellulose, which is commonly fabricated in the form of a mesh and is applied as a wound dressing. Unfortunately, the structure and the topography of current cellulose meshes are not optimal for cell growth. To enhance the surface structure and the physicochemical properties of a commercially available mesh, we coated the mesh with wood-derived cellulose nanofibrils (CNFs). Three different types of mesh coatings are proposed in this study as a skin cell carrier: positively charged cationic cellulose nanofibrils (cCNFs), negatively charged anionic cellulose nanofibrils (aCNFs), and a combination of these two materials (c+aCNFs). These cell carriers were seeded with normal human dermal fibroblasts (NHDFs) or with human adipose-derived stem cells (ADSCs) to investigate cell adhesion, spreading, morphology, and proliferation. The negatively charged aCNF coating significantly improved the proliferation of both cell types. The positively charged cCNF coating significantly enhanced the adhesion of ADSCs only. The number of NHDFs was similar on the cCNF coatings and on the noncoated pristine cellulose mesh. However, the three-dimensional (3D) structure of the cCNF coating promoted cell survival. The c+aCNF construct proved to combine benefits from both types of CNFs, which means that the c+aCNF cell carrier is a promising candidate for further application in skin tissue engineering.

• MeSH
• celulosa * MeSH
• hydrogely MeSH
• kmenové buňky MeSH
• kůže * MeSH
• lidé MeSH
• tkáňové inženýrství MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• celulosa * MeSH
• hydrogely MeSH