Allodiploid hybrid species, Aspergillus latus, belonging to section Nidulantes, is a hybrid of A. spinulosporus and an unknown species closely related to A. quadrilineatus and A. sublatus. This hybrid has often been misidentified as the species in section Nidulantes, such as A. nidulans, A. spinulosporus, A. sublatus, or other cryptic species. Aspergillus latus has not been reported in Japan as well as Asia so far. In this study, we screened 23 clinical strains identified as A. spinulosporus isolated in Japan from 2012 to 2023 and found seven A. latus strains. To characterize the A. latus strains, we conducted comprehensive phenotyping including morphological observation, whole genome sequences, and phylogenetic analysis based on calmodulin (CaM) gene. In addition, we conducted antifungal susceptibility testing for A. latus strains. As a result, the morphological characters of A. latus were more similar to those of A. spinulosporus compared to A. sublatus. However, the ascospore of A. latus differed from that of A. spinulosporus. Phylogenetic analysis revealed that different CaM alleles from the same isolate clustered separately with A. spinulosporus and A. sublatus, consistent with its hybrid origin. Furthermore, A. latus strains showed reduced susceptibility to caspofungin and amphotericin B compared to A. spinulosporus, while they were susceptible to azoles. Our results suggest that A. latus has been a causative pathogen of aspergillosis in Japan since 2013.
- MeSH
- antifungální látky farmakologie MeSH
- Aspergillus * genetika klasifikace izolace a purifikace účinky léků MeSH
- aspergilóza * mikrobiologie epidemiologie MeSH
- fylogeneze MeSH
- kalmodulin genetika MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- sekvenování celého genomu MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Japonsko MeSH
Transient receptor potential melastatin (TRPM) channels, a subfamily of the TRP superfamily, constitute a diverse group of ion channels involved in mediating crucial cellular processes like calcium homeostasis. These channels exhibit complex regulation, and one of the key regulatory mechanisms involves their interaction with calmodulin (CaM), a cytosol ubiquitous calcium-binding protein. The association between TRPM channels and CaM relies on the presence of specific CaM-binding domains in the channel structure. Upon CaM binding, the channel undergoes direct and/or allosteric structural changes and triggers down- or up-stream signaling pathways. According to current knowledge, ion channel members TRPM2, TRPM3, TRPM4, and TRPM6 are directly modulated by CaM, resulting in their activation or inhibition. This review specifically focuses on the interplay between TRPM channels and CaM and summarizes the current known effects of CaM interactions and modulations on TRPM channels in cellular physiology.
Constantly increasing attention to bioengineered proteins has led to the rapid development of new functional targets. Here we present the biophysical and functional characteristics of the newly designed CaM/AMBN-Ct fusion protein. The two-domain artificial target consists of calmodulin (CaM) and ameloblastin C-terminus (AMBN-Ct). CaM as a well-characterized calcium ions (Ca2+) binding protein offers plenty of options in terms of Ca2+ detection in biomedicine and biotechnologies. Highly negatively charged AMBN-Ct belongs to intrinsically disordered proteins (IDPs). CaM/AMBN-Ct was designed to open new ways of communication synergies between the domains with potential functional improvement. The character and function of CaM/AMBN-Ct were explored by biophysical and molecular modelling methods. Experimental studies have revealed increased stability and preserved CaM/AMBN-Ct function. The results of molecular dynamic simulations (MDs) outlined different interface patterns between the domains with potential allosteric communication within the fusion.
- MeSH
- kalmodulin chemie MeSH
- lidé MeSH
- molekulární modely MeSH
- proteiny zubní skloviny chemie metabolismus MeSH
- sekvence aminokyselin genetika MeSH
- vápník chemie MeSH
- vazba proteinů fyziologie MeSH
- vazebná místa fyziologie MeSH
- vnitřně neuspořádané proteiny chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Kinesin-like calmodulin-binding protein (KCBP) is a unique kinesin with half kinesin and half myosin, with kinesin motor domain at C-terminus and myosin tail homology region 4 (MyTH4) and band 4.1, ezrin, radixin, moesin (FERM) domains at N-terminus. The special structure endows KCBP multi-intracellular functions, including cell division, trichome morphogenesis in plants, and flagellar function in algae. However, little is known about the molecular mechanism underlying these functions. Here, we identified a molecular chaperone Hsp90 as a novel binding partner with KCBP in Dunaliella salina using a yeast two-hybrid screen. Further analysis showed that Hsp90 interacted with both the N-terminal and C-terminal of DsKCBP. Since Hsp90 was involved in the stability and proteolytic turnover of numerous proteins, whether Hsp90 regulated the degradation of DsKCBP was investigated. Our results showed that both Hsp90 and DsKCBP presented in the purified proteasome, and the interaction of DsKCBP-Hsp90 was inhibited upon Hsp90 inhibitor geldanamycin treatment. The level of DsKCBP proteins was diminished remarkably indicating that the disassociation of DsKCBP from Hsp90 accelerated the degradation of the former. Furthermore, immunofluorescence results showed that the localization of DsKCBP at basal body and flagella was disappeared by Hsp90 inhibition. The increased mRNA level of DsKCBP during flagellar assembly was not obvious by geldanamycin treatment. These data provided evidence that Hsp90 protected DsKCBP from degradation by proteasome and was involved in the role of DsKCBP in flagellar assembly.
Molecular determinants of the binding of various endogenous modulators to transient receptor potential (TRP) channels are crucial for the understanding of necessary cellular pathways, as well as new paths for rational drug designs. The aim of this study was to characterise interactions between the TRP cation channel subfamily melastatin member 4 (TRPM4) and endogenous intracellular modulators-calcium-binding proteins (calmodulin (CaM) and S100A1) and phosphatidylinositol 4, 5-bisphosphate (PIP2). We have found binding epitopes at the N- and C-termini of TRPM4 shared by CaM, S100A1 and PIP2. The binding affinities of short peptides representing the binding epitopes of N- and C-termini were measured by means of fluorescence anisotropy (FA). The importance of representative basic amino acids and their combinations from both peptides for the binding of endogenous TRPM4 modulators was proved using point alanine-scanning mutagenesis. In silico protein-protein docking of both peptides to CaM and S100A1 and extensive molecular dynamics (MD) simulations enabled the description of key stabilising interactions at the atomic level. Recently solved cryo-Electron Microscopy (EM) structures made it possible to put our findings into the context of the entire TRPM4 channel and to deduce how the binding of these endogenous modulators could allosterically affect the gating of TRPM4. Moreover, both identified binding epitopes seem to be ideally positioned to mediate the involvement of TRPM4 in higher-order hetero-multimeric complexes with important physiological functions.
- MeSH
- akvaporiny chemie metabolismus MeSH
- interakční proteinové domény a motivy * MeSH
- kalmodulin chemie metabolismus MeSH
- kationtové kanály TRPM chemie metabolismus MeSH
- kinetika MeSH
- konformace proteinů MeSH
- lidé MeSH
- molekulární modely MeSH
- multiproteinové komplexy chemie metabolismus MeSH
- peptidové fragmenty MeSH
- proteiny S100 chemie metabolismus MeSH
- sekvence aminokyselin MeSH
- vazba proteinů MeSH
- vazebná místa * MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Klíčová slova
- beauverolidy, beauveriolidy,
- MeSH
- Alzheimerova nemoc farmakoterapie patofyziologie MeSH
- amyloidní beta-protein MeSH
- ateroskleróza farmakoterapie patofyziologie MeSH
- Beauveria metabolismus MeSH
- cholesterolacyltransferasa antagonisté a inhibitory fyziologie MeSH
- cyklické peptidy terapeutické užití MeSH
- depsipeptidy * terapeutické užití MeSH
- fungální proteiny * biosyntéza MeSH
- kalmodulin antagonisté a inhibitory fyziologie MeSH
- lidé MeSH
- makrofágy patologie účinky léků MeSH
- myši MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- Publikační typ
- práce podpořená grantem MeSH
Transient receptor potential (TRPs) channels are crucial downstream targets of calcium signalling cascades. They can be modulated either by calcium itself and/or by calcium-binding proteins (CBPs). Intracellular messengers usually interact with binding domains present at the most variable TRP regions-N- and C-cytoplasmic termini. Calmodulin (CaM) is a calcium-dependent cytosolic protein serving as a modulator of most transmembrane receptors. Although CaM-binding domains are widespread within intracellular parts of TRPs, no such binding domain has been characterised at the TRP melastatin member-the transient receptor potential melastatin 6 (TRPM6) channel. Another CBP, the S100 calcium-binding protein A1 (S100A1), is also known for its modulatory activities towards receptors. S100A1 commonly shares a CaM-binding domain. Here, we present the first identified CaM and S100A1 binding sites at the N-terminal of TRPM6. We have confirmed the L520-R535 N-terminal TRPM6 domain as a shared binding site for CaM and S100A1 using biophysical and molecular modelling methods. A specific domain of basic amino acid residues (R526/R531/K532/R535) present at this TRPM6 domain has been identified as crucial to maintain non-covalent interactions with the ligands. Our data unambiguously confirm that CaM and S100A1 share the same binding domain at the TRPM6 N-terminus although the ligand-binding mechanism is different.
- MeSH
- kalmodulin chemie MeSH
- kationtové kanály TRPM chemie MeSH
- lidé MeSH
- molekulární modely * MeSH
- proteinové domény MeSH
- proteiny S100 chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Fungi under the order Ophiostomatales (Ascomycota) are known to associate with various species of bark beetles (Coleoptera: Curculionidae: Scolytinae). In addition this group of fungi contains many taxa that can impart blue-stain on sapwood and some are important tree pathogens. A recent survey that focussed on the diversity of the Ophiostomatales in the forest ecosystems of the Czech Republic and Poland uncovered four putative new species. Phylogenetic analyses of four gene regions (ITS1-5.8S-ITS2 region, ß-tubulin, calmodulin, and translation elongation factor 1-α) indicated that these four species are members of the genus Ophiostoma. All four newly described species can be distinguished from each other and from closely related species based on DNA sequence comparisons, morphological characters, growth rates, and their insect associations. Based on this study four new taxa can be circumscribed and the following names are provided: Ophiostoma pityokteinis sp. nov., Ophiostoma rufum sp. nov., Ophiostoma solheimii sp. nov., and Ophiostoma taphrorychi sp. nov. O. rufum sp. nov. is a member of the Ophiostoma piceae species complex, while O. pityokteinis sp. nov. resides in a discrete lineage within Ophiostoma s. stricto. O. taphrorychi sp. nov. together with O. distortum formed a well-supported clade in Ophiostoma s. stricto close to O. pityokteinis sp. nov. O. solheimii sp. nov. groups within a currently undefined lineage A, which also includes Ophiostoma grandicarpum and Ophiostoma microsporum. This study highlights the need for more intensive surveys that should include additional countries of Central Europe, insect vectors and host tree species in order to elucidate Ophiostoma species diversity in this region.
- MeSH
- cévnaté rostliny parazitologie MeSH
- dřevo parazitologie MeSH
- elongační faktor 1 genetika MeSH
- fylogeneze * MeSH
- kalmodulin genetika MeSH
- mezerníky ribozomální DNA genetika MeSH
- mykologické určovací techniky MeSH
- nosatcovití růst a vývoj mikrobiologie MeSH
- Ophiostoma klasifikace genetika izolace a purifikace fyziologie MeSH
- RNA ribozomální 5.8S genetika MeSH
- sekvenční analýza DNA MeSH
- tubulin genetika MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
- Polsko MeSH
The transient receptor potential channel of melastatin 4 (TRPM4) belongs to a group of large ion receptors that are involved in countless cell signalling cascades. This unique member is ubiquitously expressed in many human tissues, especially in cardiomyocytes, where it plays an important role in cardiovascular processes. Transient receptor potential channels (TRPs) are usually constituted by intracellular N- and C- termini, which serve as mediators affecting allosteric modulation of channels, resulting in the regulation of the channel function. The TRPs tails contain a number of conserved epitopes that specifically bind the intracellular modulators. Here, we identify new binding sites for the calmodulin (CaM) and S100 calcium-binding protein A1 (S100A1), located in the very distal part of the TRPM4 N terminus. We have used chemically synthesized peptides of the TRPM4, mimicking the binding epitopes, along with fluorescence methods to determine and specify CaM- and S100A1-binding sites. We have found that the ligands binding epitopes at the TRPM4 N terminus overlap, but the interacting mechanism of both complexes is probably different. The molecular models supported by data from the fluorescence method confirmed that the complexes formations are mediated by the positively charged (R139, R140, R144) and hydrophobic (L134, L138, V143) residues present at the TRPM4 N terminus-binding epitopes. The data suggest that the molecular complexes of TRPM4/CaM and TRPM4/S100A1 would lead to the modulation of the channel functions.
- MeSH
- databáze proteinů MeSH
- epitopy MeSH
- expertní systémy MeSH
- fluorescenční polarizace MeSH
- interakční proteinové domény a motivy MeSH
- kalmodulin chemie genetika metabolismus MeSH
- kationtové kanály TRPM chemie genetika metabolismus MeSH
- kinetika MeSH
- konformace proteinů MeSH
- konzervovaná sekvence MeSH
- lidé MeSH
- ligandy MeSH
- molekulární modely * MeSH
- mutace MeSH
- peptidové fragmenty chemická syntéza chemie genetika metabolismus MeSH
- proteiny S100 chemie genetika metabolismus MeSH
- rekombinantní proteiny chemie metabolismus MeSH
- sekvence aminokyselin MeSH
- simulace molekulového dockingu MeSH
- substituce aminokyselin MeSH
- vazebná místa MeSH
- výpočetní biologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Invasive fungal disease represents one of the severe complications in haematopoietic stem cell transplant recipients. We describe a case of a patient treated for relapse of chronic lymphoblastic leukaemia 6 years after HSCT. The patient was treated for invasive pulmonary aspergillosis but died 3 months later from multiple organ failures consisting of haemorrhagic necrotizing fungal pneumonia, refractory chronic hepatic graft versus host disease and cytomegalovirus hepatitis. Autopsy samples revealed histopathological evidence of fungal hyphae and an unusual Aspergillus nidulans-like species was isolated in pure culture. More precise identification was achieved by using scanning electron microscopy of ascospores and sequencing of calmodulin gene, and the isolate was subsequently re-identified as A. sublatus (section Nidulantes) and showed good in vitro susceptibility against all classes of antifungals. Commonly used ITS rDNA region and β-tubulin gene fail to discriminate A. sublatus from related pathogenic species, especially A. quadrilineatus and A. nidulans. Although this is the first case of proven IPA attributed to A. sublatus, we demonstrated that at least some previously reported infections due to A. quadrilineatus were probably caused by this cryptic species.
- MeSH
- antifungální látky aplikace a dávkování MeSH
- Aspergillus klasifikace cytologie genetika izolace a purifikace MeSH
- chronická lymfatická leukemie komplikace MeSH
- cytomegalovirové infekce komplikace diagnóza MeSH
- DNA fungální chemie genetika MeSH
- fatální výsledek MeSH
- fylogeneze MeSH
- invazivní plicní aspergilóza diagnóza farmakoterapie mikrobiologie patologie MeSH
- kalmodulin genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- mezerníky ribozomální DNA chemie genetika MeSH
- mikrobiální testy citlivosti MeSH
- mikroskopie elektronová rastrovací MeSH
- nemoc štěpu proti hostiteli komplikace diagnóza MeSH
- příjemce transplantátu MeSH
- sekvenční analýza DNA MeSH
- shluková analýza MeSH
- transplantace hematopoetických kmenových buněk škodlivé účinky MeSH
- tubulin genetika MeSH
- virová hepatitida u lidí komplikace diagnóza MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
