Increase of HCN current in SOD1-associated amyotrophic lateral sclerosis.

The clinical manifestations of sporadic amyotrophic lateral sclerosis (ALS) vary widely. However, the current classification of ALS is mainly based on clinical presentations, while the roles of electrophysiological and biomedical biomarkers remain limited. Herein, we investigated a group of patients with sporadic ALS and an ALS mouse model with superoxide dismutase 1 (SOD1)/G93A transgenes using nerve excitability tests (NET) to investigate axonal membrane properties and chemical precipitation, followed by enzyme-linked immunosorbent assay analysis to measure plasma misfolded protein levels. Six of 19 patients (31.6%) with sporadic ALS had elevated plasma misfolded SOD1 protein levels. In sporadic ALS patients, only those with elevated misfolded SOD1 protein levels showed an increased inward rectification in the current-threshold (I/V) curve and an increased threshold reduction in the hyperpolarizing threshold electrotonus (TE) in the NET study. Two familial ALS patients with SOD1 mutations also exhibited similar electrophysiological patterns of NET. For patients with sporadic ALS showing significantly increased inward rectification in the I/V curve, we noted an elevation in plasma misfolded SOD1 level, but not in total SOD1, misfolded C9orf72, or misfolded phosphorylated TDP43 levels. Computer simulations demonstrated that the aforementioned axonal excitability changes are likely associated with an increase in hyperpolarization-activated cyclic nucleotide-gated (HCN) current. In SOD1/G93A mice, NET also showed an increased inward rectification in the I/V curve, which could be reversed by a single injection of the HCN channel blocker, ZD7288. Daily treatment of SOD1/G93A mice with ZD7288 partially prevented the early motor function decline and spinal motor neuron death. In summary, sporadic ALS patients with elevated plasma misfolded SOD1 exhibited similar patterns of motor axonal excitability changes as familial ALS patients and ALS mice with mutant SOD1 genes, suggesting the existence of SOD1-associated sporadic ALS. The observed NET pattern of increased inward rectification in the I/V curve was attributable to an elevation in the HCN current in SOD1-associated ALS.

An acidophilic fungus promotes prey digestion in a carnivorous plant.

Leaves of the carnivorous sundew plants (Drosera spp.) secrete mucilage that hosts microorganisms, but whether this microbiota contributes to prey digestion is unclear. We identified the acidophilic fungus Acrodontium crateriforme as the dominant species in the mucilage microbial communities, thriving in multiple sundew species across the global range. The fungus grows and sporulates on sundew glands as its preferred acidic environment, and its presence in traps increased the prey digestion process. A. crateriforme has a reduced genome similar to other symbiotic fungi. During A. crateriforme-Drosera spatulata coexistence and digestion of prey insects, transcriptomes revealed significant gene co-option in both partners. Holobiont expression patterns during prey digestion further revealed synergistic effects in several gene families including fungal aspartic and sedolisin peptidases, facilitating prey digestion in leaves, as well as nutrient assimilation and jasmonate signalling pathway expression. This study establishes that botanical carnivory is defined by adaptations involving microbial partners and interspecies interactions.

Restoring functional TDP-43 oligomers in ALS and laminopathic cellular models through baicalein-induced reconfiguration of TDP-43 aggregates.

A group of misfolded prone-to-aggregate domains in disease-causing proteins has recently been shown to adopt unique conformations that play a role in fundamental biological processes. These processes include the formation of membrane-less sub-organelles, alternative splicing, and gene activation and silencing. The cellular responses are regulated by the conformational switching of prone-to-aggregate domains, independently of changes in RNA or protein expression levels. Given this, targeting the misfolded states of disease-causing proteins to redirect them towards their physiological conformations is emerging as an effective therapeutic strategy for diseases caused by protein misfolding. In our study, we successfully identified baicalein as a potent structure-correcting agent. Our findings demonstrate that baicalein can reconfigure existing TDP-43 aggregates into an oligomeric state both in vitro and in disease cells. This transformation effectively restores the bioactivity of misfolded TDP-43 proteins in cellular models of ALS and premature aging in progeria. Impressively, in progeria cells where defective lamin A interferes with TDP-43-mediated exon skipping, the formation of pathological TDP-43 aggregates is promoted. Baicalein, however, restores the functionality of TDP-43 and mitigates nuclear shape defects in these laminopathic cells. This establishes a connection between lamin A and TDP-43 in the context of aging. Our findings suggest that targeting physiological TDP-43 oligomers could offer a promising therapeutic avenue for treating aging-associated disorders.

Maternal Caregiving for Children Newly Diagnosed with Acute Lymphoblastic Leukemia: Traditional Chinese Mothering as the Double-Edged Sword.

AIMS AND OBJECTIVES: To explore the meaning of maternal caregiving in the Chinese culture for children newly diagnosed with acute lymphocytic leukemia (ALL). BACKGROUND: Recurrence of and death associated with ALL remain the main concerns for mothers. Mothers experience guilt and anxiety towards their child's cancer. DESIGN: Descriptive phenomenological study. METHODS: Twelve mothers were recruited from a medical centre in Central Taiwan. The mothers were primary caregivers for their child diagnosed as having ALL in the past 3 months to 1 year. Data were collected through semi-structured interviews and analysed using Colaizzi's method. RESULTS: Four main themes emerged: feeling this world crashing by knowing the diagnosis, feeling the double-edged sword of mothering, worrying about potential risks for their vulnerable child, and passing through difficulties with power of support. CONCLUSIONS: Most mothers felt this world crashing due to potential loss of their child and seeing their child's suffering. The mother was blamed for her child's cancer but was also required to shoulder all caregiving for their child. The mothers needed to compromise their lives to protect their child from potential infection. Perceived power of support helped the mothers overcome difficulties. RELEVANCE TO CLINICAL PRACTICE: Findings support that nurses encouraging mothers to tell their stories, regardless of culture, will facilitate healing. Establishing trust and providing support from nurses, physicians, psychologists and social workers will lead mothers' readiness to deal with care of their sick child. Increasing visiting time for parental support for children hospitalized in the PICU is suggested as well.

Heat-shock protein dysregulation is associated with functional and pathological TDP-43 aggregation.

Conformational disorders are involved in various neurodegenerative diseases. Reactive oxygen species (ROS) are the major contributors to neurodegenerative disease; however, ROS that affect the structural changes in misfolded disease proteins have yet to be well characterized. Here we demonstrate that the intrinsic propensity of TDP-43 to aggregate drives the assembly of TDP-43-positive stress granules and soluble toxic TDP-43 oligomers in response to a ROS insult via a disulfide crosslinking-independent mechanism. Notably, ROS-induced TDP-43 protein assembly correlates with the dynamics of certain TDP-43-associated chaperones. The heat-shock protein (HSP)-90 inhibitor 17-AAG prevents ROS-induced TDP-43 aggregation, alters the type of TDP-43 multimers and reduces the severity of pathological TDP-43 inclusions. In summary, our study suggests that a common mechanism could be involved in the pathogenesis of conformational diseases that result from HSP dysregulation.

The formation stability, hydrolytic behavior, mass spectrometry, DFT study, and luminescence properties of trivalent lanthanide complexes of H2ODO2A.

The trivalent lanthanide complex formation constants (log K(f)) of the macrocyclic ligand H(2)ODO2A (4,10-dicarboxymethyl-1-oxa-4,7,10-triazacyclododecane) have been determined by pH titration techniques to be in the range 10.84-12.62 which increase with increasing lanthanide atomic number, and are smaller than those of the corresponding H(2)DO2A (1,7-dicarboxylmethyl-1,4,7,10-tetraazacyclododecane) complexes. The equilibrium formation of the dinuclear hydrolysis species, e.g. Ln(2)(ODO2A)(2)(µ-OH)(+) and Ln(2)(ODO2A)(2)(µ-OH)(2), dominates over the mononuclear species, e.g. LnODO2A(OH) and LnODO2A(OH)(2)(-). Mass spectrometry confirmed the presence of [Eu(ODO2A)](+), [Eu(ODO2A)(OH)+H](+), [Eu(2)(ODO2A)(2)(OH(2))(2)+H](+), [Eu(ODO2A)(OH)(2)](-) and [Eu(2)(ODO2A)(2)(OH(2))(3)](-) species at pH > 7. Density function theory (DFT) calculated structures of the EuODO2A(H(2)O)(3)(+) and EuDO2A(H(2)O)(3)(+) complexes indicate that three inner-sphere coordinated water molecules are arranged in a meridional configuration, i.e. the 3 water molecules are on the same plane perpendicular to that of the basal N(3)O or N(4) atoms. However, luminescence lifetime studies reveal that the EuODO2A(+) and TbODO2A(+) complexes have 4.1 and 2.9 inner-sphere coordinated water molecules, respectively, indicating that other equilibrium species are also present for the EuODO2A(+) complex. The respective emission spectral intensities and lifetimes at 615 nm (λ(ex) = 395 nm) and 544 nm (λ(ex) = 369 nm) of the EuODO2A(+) and TbODO2A(+) complexes increase with increasing pH, consistent with the formation of µ-OH-bridged dinuclear species at higher pH. Additional DFT calculations show that each Y(iii) ion is 8-coordinated in the three possible cis-[Y(2)(ODO2A)(2)(µ-OH)(H(2)O)(2)](+), trans-[Y(2)(ODO2A)(2)(µ-OH)(H(2)O)(2)](+) and [Y(2)(ODO2A)(2)(µ-OH)(2)] dinuclear complex structures. The first and the second include 6-coordination by the ligand ODO2A(2-), one by the bridged µ-OH ion and one by a water molecule. The third includes 6-coordination by the ligand ODO2A(2-) and two by the bridged µ-OH ions. The two inner-sphere coordinated water molecules in the cis- and trans-[Y(2)(ODO2A)(2)(µ-OH)(H(2)O)(2)](+) dinuclear complexes are in a staggered conformation with torsional angles of 82.21° and 148.54°, respectively.

The self-interaction of native TDP-43 C terminus inhibits its degradation and contributes to early proteinopathies.

The degraded, misfolded C terminus of TAR DNA-binding protein-43 is associated with a wide spectrum of neurodegenerative diseases, particularly frontotemporal lobar degeneration with ubiquitin-positive inclusions and amyotrophic lateral sclerosis. However, the precise mechanism of pathological cleavage of the TAR DNA-binding protein-43 remains unknown. Here we show that the TAR DNA-binding protein-43 C-terminal protein physically interacts with itself or with the cellular-folded yeast prion domain of Sup35 forming dynamic aggregates. This prion-like nature governs known cellular functions of the TAR DNA-binding protein-43, including subcellular localisation and exon skipping of the cystic fibrosis transmembrane conductance regulator. Significantly, mutants with a failure to engage in prion-like interactions are processed into an ~24-kDa C-terminal fragment of the TAR DNA-binding protein-43. The estimated cleavage site of degraded TAR DNA-binding protein-43 fragments corresponds to the pathological cleavage site identified in patients with the TAR DNA-binding protein-43 proteinopathies. Consistently, epigallocatechin gallate constrains prion-like interactions, attenuating pathological-like degradation. Thus, the native folding of TAR DNA-binding protein-43 C terminus acts as a guardian of pathogenesis, which is directly associated with loss-of-function.

TDP-43: an emerging new player in neurodegenerative diseases.

Until a couple of years ago, TAR-DNA-binding protein-43 (TDP-43) was a relatively unknown nuclear protein implicated in transcriptional repression and splicing. Since 2006, when the protein was reported to be present in inclusions in the neurons and/or glial cells of a range of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration with ubiquitin-positive, tau- and alpha-synuclein-negative inclusions (FTLD-U) and Alzheimer's disease (AD), many reports on the medical aspects of TDP-43 have been published. Here, we summarize the current literature on TDP-43, focusing on recent studies that provide clues to the function of TDP-43. Using this information and database analysis, we also suggest a molecular and cellular model for possible events in normal and diseased neurons in relation to the emerging importance of the function and dysfunction of this protein as a target for basic as well as translational research.

TDP-43 overexpression enhances exon 7 inclusion during the survival of motor neuron pre-mRNA splicing.

TDP-43 is a highly conserved, 43-kDa RNA-binding protein implicated to play a role in transcription repression, nuclear organization, and alternative splicing. More recently, this factor has been identified as the major disease protein of several neurodegenerative diseases, including frontotemporal lobar degeneration with ubiquitin-positive inclusions and amyotrophic lateral sclerosis. For the splicing activity, the factor has been shown to be mainly an exon-skipping promoter. In this study using the survival of motor neuron (SMN) minigenes as the reporters in transfection assay, we show for the first time that TDP-43 could also act as an exon-inclusion factor. Furthermore, both RNA-recognition motif domains are required for its ability to enhance the SMN2 exon 7 inclusion. Combined protein-immunoprecipitation and RNA-immunoprecipitation experiments also suggested that this exon inclusion activity might be mediated by multimeric complex(es) consisting of this protein interacting with other splicing factors, including Htra2-beta1. Our data further evidence TDP-43 as a multifunctional RNA-binding protein for a diverse set of cellular activities.

TDP-43, the signature protein of FTLD-U, is a neuronal activity-responsive factor.

TDP-43, recently identified as a signature protein of the pathogenic inclusions in the brains cells of frontotemporal lobar degeneration patients, is a 43 kDa RNA-binding protein. It has been known mainly as a nuclear factor capable of repressing transcription and promoting exon exclusion. TDP-43 also forms distinct nuclear substructures linking different types of nuclear bodies. In this study, we provide the first evidence supporting TDP-43 as a neuronal activity-responsive factor in the dendrites of hippocampal neurons. In particular, TDP-43 resides in the somatodendrites mainly in the form of RNA granules colocalized with the post-synaptic protein PSD-95. These granules also contain RNAs including at least the beta-actin mRNA and CaMKIIalpha mRNA. Furthermore, TDP-43 is localized in the dendritic processing (P) body and it behaves as a translational repressor in an in vitro assay. Related to this, repetitive stimuli by KCl greatly enhance the colocalization of TDP-43 granules with FMRP and Staufen 1, two RNA-binding proteins known to regulate mRNA transport and local translation in neurons. These data together suggest that TDP-43 is a neuronal activity-responsive factor functioning in the regulation of neuronal plasticity, the impairment of which would lead to the development of certain forms of neurodegenerative diseases including frontotemporal lobar degeneration.

A post-synaptic scaffold at the origin of the animal kingdom.

BACKGROUND: The evolution of complex sub-cellular structures such as the synapse requires the assembly of multiple proteins, each conferring added functionality to the integrated structure. Tracking the early evolution of synapses has not been possible without genomic information from the earliest branching animals. As the closest extant relatives to the Eumetazoa, Porifera (sponges) represent a pivotal group for understanding the evolution of nervous systems, because sponges lack neurons with clearly recognizable synapses, in contrast to eumetazoan animals. METHODOLOGY/PRINCIPAL FINDINGS: We show that the genome of the demosponge Amphimedon queenslandica possesses a nearly complete set of post-synaptic protein homologs whose conserved interaction motifs suggest assembly into a complex structure. In the critical synaptic scaffold gene, dlg, residues that make hydrogen bonds and van der Waals interactions with the PDZ ligand are 100% conserved between sponge and human, as is the motif organization of the scaffolds. Expression in Amphimedon of multiple post-synaptic gene homologs in larval flask cells further supports the existence of an assembled structure. Among the few post-synaptic genes absent from Amphimedon, but present in Eumetazoa, are receptor genes including the entire ionotropic glutamate receptor family. CONCLUSIONS/SIGNIFICANCE: Highly conserved protein interaction motifs and co-expression in sponges of multiple proteins whose homologs interact in eumetazoan synapses indicate that a complex protein scaffold was present at the origin of animals, perhaps predating nervous systems. A relatively small number of crucial innovations to this pre-existing structure may represent the founding changes that led to a post-synaptic element.

Actin-based modeling of a transcriptionally competent nuclear substructure induced by transcription inhibition.

During transcription inactivation, the nuclear bodies in the mammalian cells often undergo reorganization. In particular, the interchromatin granule clusters, or IGCs, become colocalized with RNA polymerase II (RNAP II) upon treatment with transcription inhibitors. This colocalization has also been observed in untreated but transcriptionally inactive cells. We report here that the reorganized IGC domains are unique substructure consisting of outer shells made of SC35, ERK2, SF2/ASF, and actin. The apparently hollow holes of these domains contain clusters of RNAP II, mostly phosphorylated, and the splicing regulator SMN. This class of complexes are also the sites where prominent transcription activities are detected once the inhibitors are removed. Furthermore, actin polymerization is required for reorganization of the IGCs. In connection with this, immunoprecipitation and immunostaining experiments showed that nuclear actin is associated with IGCs and the reorganized IGC domains. The study thus provides further evidence for the existence of an actin-based nuclear skeleton structure in association with the dynamic reorganization processes in the nucleus. Overall, our data suggest that mammalian cells have adapted to utilize the reorganized, uniquely shaped IGC domains as the temporary storage sites of RNAP II transcription machineries in response to certain transient states of transcription inactivation.

Gitelman syndrome: report of three cases and literature review.

Gitelman syndrome (GS) is a rare autosomal recessive, inherited renal tubular disorder. Herein, we report three cases of GS, one sporadic case and two siblings. They have typical laboratory findings, including hypokalemia, metabolic alkalosis, hypomagnesemia, and hypocalciuria. All of them were treated with oral potassium and magnesium supplements. They received regular pediatric clinic follow-up to check electrolytes and monitor development. These three cases reminded us that doctors should be alert to unexplained hypokalemia, which is usually the initial presentation of GS.

Structural diversity and functional implications of the eukaryotic TDP gene family.

TDP-43 is an RNA-binding protein that functions in mammalian cells in transcriptional repression and exon skipping. The gene encoding TDP-43 (HGMW-approved gene symbol TARDBP) is conserved in human, mouse, Drosophila melanogaster, and Caenorhabditis elegans. Sequence comparison of the coding regions of the TDP genes among the four taxa reveals an extraordinarily low rate of sequence divergence, suggesting that the TDP genes carry out essential functions in these organisms. With DNA transfection assay, we have established the importance of the glycine-rich domain for the exon-skipping activity of TDP-43. Both human and mouse TDP genes belong to a gene family that also consists of a number of processed pseudogenes. Interestingly, combined database analysis and cDNA cloning have demonstrated that the primary transcript of the mammalian TDP genes undergoes alternative splicing to generate 11 mRNAs, including the one encoding TDP-43. Eight of the 11 splicing events involved the use of four each of the 5'-donor and 3'-acceptor sites, all of which reside within the last exon of the TDP-43 mRNA. The existence of multispliced isoforms of TDP-encoded proteins provides further support for the functional complexity of the eukaryotic TDP genes.

Higher order arrangement of the eukaryotic nuclear bodies.

The nuclei of eukaryotic cells consist of discrete substructures. These substructures include the nuclear bodies, which have been implicated in a number of biological processes such as transcription and splicing. However, for most nuclear bodies, the details of involvement in these processes in relation to their three-dimensional distributions in the nucleus are still unclear. Through the analysis of TDP, a protein functional in both transcriptional repression and alternative splicing, we have identified a new category of nuclear bodies within which the TDP molecules reside. Remarkably, the TDP bodies (TBs) colocalize or overlap with several different types of nuclear bodies previously suggested to function in transcription or splicing. Of these nuclear bodies, the Gemini of coiled bodies (GEM) seems to associate with TB through the interaction between survival motor neuron (SMN) protein and TDP. Furthermore, TB sometimes appears to be the bridge of two or more of these other nuclear bodies. Our data suggest the existence of a hierarchy and possibly functional arrangement of the nuclear bodies within the eukaryotic nuclei.