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Objectives: To describe how patients with Parkinson’s disease (PD) and care partners choose to share or withhold information from clinicians. Methods: This is a qualitative, descriptive study nested within a multisite, randomized clinical trial of outpatient palliative care compared to standard neurologic care for PD. Interviews with patients (n = 30) and care partners (n = 30) explored experiences communicating with neurology clinicians. Thematic analyses identified themes relevant to patient-care partner-clinician communication. Results: There were four themes relevant to sharing and/or withholding information from clinicians: (1) Suppressing Concerns During Visits, (2) Care Partner Awareness of Patients’ Communication Barriers due to Cognitive Impairment, (3) Limited Sharing of Sensitive or Intimate Issues by Patients and Care Partners, and (4) Patient and Care Partner Suggestions to Overcome ‘Holding Back’. Conclusion: Limitations to communication between patients, care partners, and clinicians should be acknowledged and recognized in routine Parkinson’s disease care to foster accurate disclosure of unmet palliative care and other needs. Triadic communication strategies may help patients and care partners talk about unmet palliative care needs. Innovation: By recognizing that cognitive impairment and sensitive topics can be barriers, clinicians can adjust or adopt targeted communication strategies for identifying and discussing care needs.

Abstract Background Veterans who need post-acute home health care (HHC) are at risk for adverse outcomes and unmet social needs. Veterans’ social needs could be identified and met by community-based HHC clinicians due to their unique perspective from the home environment, acuity of Veterans they serve, and access to Veterans receiving community care. To understand these needs, we explored clinician, Veteran, and care partner perspectives to understand Veterans’ social needs during the transition from hospital to home with skilled HHC. Methods Qualitative data were collected through individual interviews with Veterans Health Administration (VHA) inpatient & community HHC clinicians, Veterans, and care partners who have significant roles facilitating Veterans’ hospital to home with HHC transition. To inform implementation of a care coordination quality improvement intervention, participants were asked about VHA and HHC care coordination and Veterans’ social needs during these transitions. Interviews were recorded, transcribed, and analyzed inductively using thematic analysis and results were organized deductively according to relevant transitional care domains (Discharge Planning, Transition to Home, and HHC Delivery). Results We conducted 35 interviews at 4 VHA Medical Centers located in Western, Midwestern, and Southern U.S. regions during March 2021 through July 2022. We organized results by the three care transition domains and related themes by VHA, HHC, or Veteran/care partner perspective. Our themes included (1) how social needs affected access to HHC, (2) the need for social needs screening during hospitalization, (3) delays in HHC for Veterans discharged from community hospitals, and (4) a need for closed-loop communication between VHA and HHC to report social needs. Conclusions HHC is an underexplored space for Veterans social needs detection. While this research is preliminary, we recommend two steps forward from this work: (1) develop closed-loop communication and education pathways with HHC and (2) develop a partnership to integrate a social risk screener into HHC pathways.

Abstract Background The Veterans Affairs (VA) Healthcare System Community Hospital Transitions Program (CHTP) was implemented as a nurse-led intervention to reduce barriers that patients experience when transitioning from community hospitals to VA primary care settings. A previous analysis indicated that veterans who enrolled in CHTP received timely follow-up care and communications that improved care coordination, but did not examine cost implications for the VA. Methods A budget impact analysis used the VA (payer) perspective. CHTP implementation team members and study records identified key resources required to initially implement and run the CHTP. Statistical analysis of program participants and matched controls at two study sites was used to estimate incremental VA primary care costs per veteran. Using combined program implementation, operations, and healthcare cost estimates to guide key model assumptions, overall CHTP costs were estimated for a 5-year time horizon, including a discount rate of 3%, annual inflation of 2.5%, and a sensitivity analysis that considered two options for staffing the program at VA Medical Center (VAMC) sites. Results Implementation at two VAMCs required 3 months, including central program support and site-level onboarding, with costs of $34,094 (range: $25,355–$51,602), which included direct and indirect resource costs of personnel time, materials, space, and equipment. Subsequent annual costs to run the program at each site depended heavily on the staffing mix and caseload of veterans, with a baseline estimate of $193,802 to $264,868. Patients enrolled in CHTP had post-hospitalization VA primary care costs that were higher than matched controls. Over 5 years, CHTP sites staffed to serve 25–30 veterans per full-time equivalent transition team member per month had an estimated budget impact of $625 per veteran served if the transitional team included a medical social worker to support veterans with more social behavioral needs and less complex medical cases or $815 per veteran if nurses served all cases. Conclusions Evidence-based care coordination programs that support patients’ return to VA primary care after a community hospital stay are feasible to implement and run. Further, flexibility in staffing this type of program is increasingly relevant as the VA and other healthcare systems consider methods to reduce provider burnout, optimize staffing, reduce costs, and address other staffing challenges while improving patient care.

Abstract Background Empirical evidence supports the use of structured goals of care conversations and documentation of life-sustaining treatment (LST) preferences in durable, accessible, and actionable orders to improve the care for people living with serious illness. As the largest integrated healthcare system in the USA, the Veterans Health Administration (VA) provides an excellent environment to test implementation strategies that promote this evidence-based practice. The Preferences Elicited and Respected for Seriously Ill Veterans through Enhanced Decision-Making (PERSIVED) program seeks to improve care outcomes for seriously ill Veterans by supporting efforts to conduct goals of care conversations, systematically document LST preferences, and ensure timely and accurate communication about preferences across VA and non-VA settings. Methods PERSIVED encompasses two separate but related implementation projects that support the same evidence-based practice. Project 1 will enroll 12 VA Home Based Primary Care (HBPC) programs and Project 2 will enroll six VA Community Nursing Home (CNH) programs. Both projects begin with a pre-implementation phase during which data from diverse stakeholders are gathered to identify barriers and facilitators to adoption of the LST evidence-based practice. This baseline assessment is used to tailor quality improvement activities using audit with feedback and implementation facilitation during the implementation phase. Site champions serve as the lynchpin between the PERSIVED project team and site personnel. PERSIVED teams support site champions through monthly coaching sessions. At the end of implementation, baseline site process maps are updated to reflect new steps and procedures to ensure timely conversations and documentation of treatment preferences. During the sustainability phase, intense engagement with champions ends, at which point champions work independently to maintain and improve processes and outcomes. Ongoing process evaluation, guided by the RE-AIM framework, is used to monitor Reach, Adoption, Implementation, and Maintenance outcomes. Effectiveness will be assessed using several endorsed clinical metrics for seriously ill populations. Discussion The PERSIVED program aims to prevent potentially burdensome LSTs by consistently eliciting and documenting values, goals, and treatment preferences of seriously ill Veterans. Working with clinical operational partners, we will apply our findings to HBPC and CNH programs throughout the national VA healthcare system during a future scale-out period.

Introduction: Implementation science frameworks have helped advance translation of research to practice. They have been widely used for planning and post-hoc evaluation, but seldom to inform and guide mid-course adjustments to intervention and implementation strategies.Materials and Methods: This study developed an innovative methodology using the RE-AIM framework and related tools to guide mid-course assessments and adaptations across five diverse health services improvement projects in the Veterans Health Administration (VA). Using a semi-structured guide, project team members were asked to assess the importance of and progress on each RE-AIM dimension (i.e., reach, effectiveness, adoption, implementation, maintenance) at the current phase of their project. Based on these ratings, each team identified one or two RE-AIM dimensions for focused attention. Teams developed proximal goals and implementation strategies to improve progress on their selected dimension(s). A follow-up meeting with each team occurred approximately 6 weeks after the goal setting meeting to evaluate the usefulness of the iterative process. Results were evaluated using both descriptive quantitative analyses and qualitative assessments from interviews and meeting notes.Results: A median of seven team members participated in the two meetings. Qualitative and descriptive data revealed that the process was feasible, understandable and useful to teams in adjusting their interventions and implementation strategies. The RE-AIM dimensions identified as most important were adoption and effectiveness, and the dimension that had the largest gap between importance and rated progress was reach. The dimensions most frequently selected for improvement were reach and adoption. Examples of action plans were summarizing stakeholder interviews for leadership, revising exclusion criteria, and conducting in-service trainings. Follow-up meetings indicated that teams found the process very useful and were able to implement the action plans they set.Discussion: The iterative use of RE-AIM to support adjustments during project implementation proved feasible and useful across diverse projects in the VA setting. Building on this and related examples, future research should replicate these findings and further develop the methodology, as well as explore the optimal frequency and timing for these iterative applications of RE-AIM. More generally, greater focus on more rapid and iterative use of implementation science frameworks is encouraged to facilitate successful translation of research to practice.

BackgroundMany health outcomes and implementation science studies have demonstrated the importance of tailoring evidence-based care interventions to local context to improve fit. By adapting to local culture, history, resources, characteristics, and priorities, interventions are more likely to lead to improved outcomes. However, it is unclear how best to adapt evidence-based programs and promising innovations. There are few guides or examples of how to best categorize or assess health-care adaptations, and even fewer that are brief and practical for use by non-researchers.Materials and methodsThis study describes the importance and potential of assessing adaptations before, during, and after the implementation of health systems interventions. We present a promising multilevel and multimethod approach developed and being applied across four different health systems interventions. Finally, we discuss implications and opportunities for future research.ResultsThe four case studies are diverse in the conditions addressed, interventions, and implementation strategies. They include two nurse coordinator-based transition of care interventions, a data and training-driven multimodal pain management project, and a cardiovascular patient-reported outcomes project, all of which are using audit and feedback. We used the same modified adaptation framework to document changes made to the interventions and implementation strategies. To create the modified framework, we started with the adaptation and modification model developed by Stirman and colleagues and expanded it by adding concepts from the RE-AIM framework. Our assessments address the intuitive domains of Who, How, When, What, and Why to classify and organize adaptations. For each case study, we discuss how the modified framework was operationalized, the multiple methods used to collect data, results to date and approaches utilized for data analysis. These methods include a real-time tracking system and structured interviews at key times during the intervention. We provide descriptive data on the types and categories of adaptations made and discuss lessons learned.ConclusionThe multimethod approaches demonstrate utility across diverse health systems interventions. The modified adaptations model adequately captures adaptations across the various projects and content areas. We recommend systematic documentation of adaptations in future clinical and public health research and have made our assessment materials publicly available.

The interplay between metal ion binding and the activity of thiol proteins, particularly within the protein disulfide isomerase family, remains an area of active investigation due to the critical role that these proteins play in many vital processes. This research investigates the interaction between recombinant human PDIA1 and zinc ions, focusing on the subsequent implications for PDIA1's conformational stability and enzymatic activity. Employing isothermal titration calorimetry and differential scanning calorimetry, we systematically compared the zinc binding capabilities of both oxidized and reduced forms of PDIA1 and assessed the structural consequences of this interaction. Our results demonstrate that PDIA1 can bind zinc both in reduced and oxidized states, but with significantly different stoichiometry and more pronounced conformational effects in the reduced form of PDIA1. Furthermore, zinc binding was observed to inhibit the catalytic activity of reduced-PDIA1, likely due to induced alterations in its conformation. These findings unveil a potential regulatory mechanism in PDIA1, wherein metal ion binding under reductive conditions modulates its activity. Our study highlights the potential role of zinc in regulating the catalytic function of PDIA1 through conformational modulation, suggesting a nuanced interplay between metal binding and protein stability in the broader context of cellular redox regulation.

Dysregulation of intraocular pressure (IOP) is one of the main risk factors for glaucoma. γ-synuclein is a member of the synuclein family of widely expressed synaptic proteins within the central nervous system that are implicated in certain types of neurodegeneration. γ-synuclein expression and localization changes in the retina and optic nerve of patients with glaucoma. However, the mechanisms by which γ-synuclein could contribute to glaucoma are poorly understood. We assessed the presence of autoantibodies to γ-synuclein in the blood serum of patients with primary open-angle glaucoma (POAG) by immunoblotting. A positive reaction was detected for five out of 25 patients (20%) with POAG. Autoantibodies to γ-synuclein were not detected in a group of patients without glaucoma. We studied the dynamics of IOP in response to IOP regulators in knockout mice (γ-KO) to understand a possible link between γ-synuclein dysfunction and glaucoma-related pathophysiological changes. The most prominent decrease of IOP in γ-KO mice was observed after the instillation of 1% phenylephrine and 10% dopamine. The total protein concentration in tear fluid of γ-KO mice was approximately two times higher than that of wild-type mice, and the activity of neurodegeneration-linked protein α2-macroglobulin was reduced. Therefore, γ-synuclein dysfunction contributes to pathological processes in glaucoma, including dysregulation of IOP.

Effectively presenting epitopes on immunogens, in order to raise conformationally selective antibodies through active immunization, is a central problem in treating protein misfolding diseases, particularly neurodegenerative diseases such as Alzheimer's disease or Parkinson's disease. We seek to selectively target conformations enriched in toxic, oligomeric propagating species while sparing the healthy forms of the protein that are often more abundant. To this end, we computationally modeled scaffolded epitopes in cyclic peptides by inserting/deleting a variable number of flanking glycines ("glycindels") to best mimic a misfolding-specific conformation of an epitope of α-synuclein enriched in the oligomer ensemble, as characterized by a region most readily disordered and solvent-exposed in a stressed, partially denatured protofibril. We screen and rank the cyclic peptide scaffolds of α-synuclein in silico based on their ensemble overlap properties with the fibril, oligomer-model and isolated monomer ensembles. We present experimental data of seeded aggregation that support nucleation rates consistent with computationally predicted cyclic peptide conformational similarity. We also introduce a method for screening against structured off-pathway targets in the human proteome by selecting scaffolds with minimal conformational similarity between their epitope and the same solvent-exposed primary sequence in structured human proteins. Different cyclic peptide scaffolds with variable numbers of glycines are predicted computationally to have markedly different conformational ensembles. Ensemble comparison and overlap were quantified by the Jensen-Shannon divergence and a new measure introduced here, the embedding depth, which determines the extent to which a given ensemble is subsumed by another ensemble and which may be a more useful measure in developing immunogens that confer conformational selectivity to an antibody.

S100A1 is a member of the S100 family of small ubiquitous Ca 2+ -binding proteins, which participates in the regulation of cell differentiation, motility, and survival. It exists as homo- or heterodimers. S100A1 has also been shown to bind Zn 2+ , but the molecular mechanisms of this binding are not yet known. In this work, using ESI-MS and ITC, we demonstrate that S100A1 can coordinate 4 zinc ions per monomer, with two high affinity (K D ~4 and 770 nm) and two low affinity sites. Using competitive binding experiments between Ca 2+ and Zn 2+ and QM/MM molecular modeling we conclude that Zn 2+ high affinity sites are located in the EF-hand motifs of S100A1. In addition, two lower affinity sites can bind Zn 2+ even when the EF-hands are saturated by Ca 2+ , resulting in a 2Ca 2+ :S100A1:2Zn 2+ conformer. Finally, we show that, in contrast to calcium, an excess of Zn 2+ produces a destabilizing effect on S100A1 structure and leads to its aggregation. We also determined a higher affinity to Ca 2+ (K D ~0.16 and 24 μm) than was previously reported for S100A1, which would allow this protein to function as a Ca 2+ /Zn 2+ -sensor both inside and outside cells, participating in diverse signaling pathways under normal and pathological conditions.

Aggregated forms of α-synuclein are core components of pathohistological inclusions known as Lewy bodies in substantia nigra (SN) neurons of patients with Parkinson's disease (PD). The role of α-synuclein in selective loss of SN dopaminergic neurons (DNs) in PD is studied in mice knocked out in the α-synuclein gene. The new mouse strain delta flox KO with a constitutive knockout of the α-synuclein gene models the end point of in vivo deletion of the α-synuclein gene in mice with a conditional knockout and has no foreign sequence in the modified genomic locus, thus differing from all other α-synuclein knockout mouse strains. The effect of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), which is used to model PD, was compared between delta flox KO mice and mice of the well-known α-synuclein knockout strain AbKO. Subchronic MPTP administration, which models early PD, was found to reduce the dopamine content and to change the ratio of dopamine metabolites in the striatum to the same levels in delta flox KO, АbKO, and wild-type mice. Overt locomotor defects were not observed after MPTP treatment, but gait testing in a CatWalk XT (Noldus) system revealed identical gait deviations in mice of the two strains and control wild-type mice. Based on the findings, a similar mechanism of neurotoxic damage to DNs was assumed for delta flox KO and AbKO mice.

Transactive response DNA and RNA binding protein 43 kDa (TDP-43) is a highly conserved heterogeneous nuclear ribonucleoprotein (hnRNP), which is involved in several steps of protein production including transcription and splicing. Its aggregates are frequently observed in motor neurons from amyotrophic lateral sclerosis patients and in the most common variant of frontotemporal lobar degeneration. Recently it was shown that TDP-43 is able to bind Zn 2+ by its RRM domain. In this work, we have investigated Zn 2+ binding to a short peptide 256-264 from C-terminus of RRM2 domain using isothermal titration calorimetry, electrospray ionization mass spectrometry, QM/MM simulations, and NMR spectroscopy. We have found that this peptide is able to bind zinc ions with a K a equal to 1.6 × 10 5 M -1 . Our findings suggest the existence of a zinc binding site in the C-terminal region of RRM2 domain. Together with the existing structure of the RRM2 domain of TDP-43 we propose a model of its complex with Zn 2+ which illustrates how zinc might regulate DNA/RNA binding.

The etiology and pathogenesis of Parkinson's disease (PD) are tightly linked to the gain-of-function of α-synuclein. However, gradual accumulation of α-synuclein aggregates in dopaminergic neurons of substantia nigra pars compacta (SNpc) leads to the depletion of the functional pool of soluble α-synuclein, and therefore, creates loss-of-function conditions, particularly in presynaptic terminals of these neurons. Studies of how this late-onset depletion of a protein involved in many important steps of neurotransmission contributes to PD progression and particularly, to worsening the nigrostriatal pathology at late stages of the disease are limited and obtained data, are controversial. Recently, we produced a mouse line for conditional knockout of the gene encoding α-synuclein, and here we used its tamoxifen-inducible pan-neuronal inactivation to study consequences of the adult-onset (from the age of 6 months) and late-onset (from the age of 12 months) α-synuclein depletion to the nigrostriatal system. No significant changes of animal balance/coordination, the number of dopaminergic neurons in the SNpc and the content of dopamine and its metabolites in the striatum were observed after adult-onset α-synuclein depletion, but in aging (18-month-old) late-onset depleted mice we found a significant reduction of major dopamine metabolites without changes to the content of dopamine itself. Our data suggest that this might be caused, at least partially, by reduced expression of aldehyde dehydrogenase ALDH1a1 and could lead to the accumulation of toxic intermediates of dopamine catabolism. By extrapolating our findings to a potential clinical situation, we suggest that therapeutic downregulation of α-synuclein expression in PD patients is a generally safe option as it should not cause adverse side effects on the functionality of their nigrostriatal system. However, if started in aged patients, this type of therapy might trigger slight functional changes of the nigrostriatal system with potentially unwanted additive effect to already existing pathology., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Multiple clinical and experimental evidences suggest that amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are members of a disease continuum. Pathological inclusions of fused in sarcoma (FUS) protein have been observed in subsets of patients with these diseases but their anatomical distribution is different for two diseases. These structures are present in motor neurons in ALS cases but in cortical neurons in FTLD cases. Expression of a C-terminally truncated form of human FUS causes an early onset and progressive motor neuron pathology in transgenic mice but only when these neurons express a certain level of this protein. Severe motor dysfunction and early lethality of mice with expression above this level prevent their use for studies of FTLD-related pathology caused by expression of this form of FUS. In the present study, we used another line of mice expressing the same protein but not developing any signs of motor system dysfunction due to substantially lower level of transgene expression in motor neurons. In a set of tests 5-month old mice displayed certain behavioural abnormalities, including increased impulsivity, decreased anxiety and compromised social interaction, which recapitulate behaviour characteristics typically seen in FTLD patients., (© 2019 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.)

Tau is an intrinsically disordered microtubule-associated protein that is implicated in several neurodegenerative disorders called tauopathies. In these diseases, Tau is found in the form of intracellular inclusions that consist of aggregated paired helical filaments (PHFs) in neurons. Given the importance of this irreversible PHF formation in neurodegenerative disease, Tau aggregation has been extensively studied. Several different factors, such as mutations or post translational modifications, have been shown to influence the formation of late-stage non-reversible Tau aggregates. It was recently shown that zinc ions accelerated heparin-induced oligomerization of Tau constructs. Indeed, in vitro studies of PHFs have usually been performed in the presence of additional co-factors, such as heparin, in order to accelerate their formation. Using turbidimetry, we investigated the impact of zinc ions on Tau in the absence of heparin and found that zinc is able to induce a temperature-dependent reversible oligomerization of Tau. The obtained oligomers were not amyloid-like and dissociated instantly following zinc chelation or a temperature decrease. Finally, a combination of isothermal titration calorimetry and dynamic light scattering experiments showed zinc binding to a high-affinity binding site and three low-affinity sites on Tau, accompanied by a change in Tau folding. Altogether, our findings stress the importance of zinc in Tau oligomerization. This newly identified Zn-induced oligomerization mechanism may be a part of a pathway different of and concurrent to Tau aggregation cascade leading to PHF formation., (Copyright © 2018. Published by Elsevier Ltd.)

Neuronal calcium sensor-1 (NCS-1) protein is abundantly expressed in the central nervous system and retinal neurons, where it regulates many vital processes such as synaptic transmission. It coordinates three calcium ions by EF-hands 2-4, thereby transducing Ca 2+ signals to a wide range of protein targets, including G protein-coupled receptors and their kinases. Here, we demonstrate that NCS-1 also has Zn 2+ -binding sites, which affect its structural and functional properties upon filling. Fluorescence and circular dichroism experiments reveal the impact of Zn 2+ binding on NCS-1 secondary and tertiary structure. According to atomic absorption spectroscopy and isothermal titration calorimetry studies, apo-NCS-1 has two high-affinity (4 × 10 6 M -1 ) and one low-affinity (2 × 10 5 M -1 ) Zn 2+ -binding sites, whereas Mg 2+ -loaded and Ca 2+ -loaded forms (which dominate under physiological conditions) bind two zinc ions with submicromolar affinity. Metal competition analysis and circular dichroism studies suggest that Zn 2+ -binding sites of apo- and Mg 2+ -loaded NCS-1 overlap with functional EF-hands of the protein. Consistently, high Zn 2+ concentrations displace Mg 2+ from the EF-hands and decrease the stoichiometry of Ca 2+ binding. Meanwhile, one of the EF-hands of Zn 2+ -saturated NCS-1 exhibits a 14-fold higher calcium affinity, which increases the overall calcium sensitivity of the protein. Based on QM/MM molecular dynamics simulations, Zn 2+ binding to Ca 2+ -loaded NCS-1 could occur at EF-hands 2 and 4. The high-affinity zinc binding increases the thermal stability of Ca 2+ -free NCS-1 and favours the interaction of its Ca 2+ -loaded form with target proteins, such as dopamine receptor D2R and GRK1. In contrast, low-affinity zinc binding promotes NCS-1 aggregation accompanied by the formation of twisted rope-like structures. Altogether, our findings suggest a complex interplay between magnesium, calcium and zinc binding to NCS-1, leading to the appearance of multiple conformations of the protein, in turn modulating its functional status.

Glioblastoma is the most frequent and aggressive primary brain tumor in adults. Recently, a growing number of studies have shown that denaturation profile of plasma samples obtained by differential scanning calorimetry (DSC) can represent a signature of a disease. In this study, we analyzed for the first time the DSC denaturation profiles of the plasma from patients with recurrent glioblastoma (n=17). Comparison to the one of healthy individuals (n=10) and to already described profiles in others cancer showed clear differences suggesting that this DSC profile may constitute a signature of glioblastoma. Parameters extracted from these profiles were used for cluster analysis which revealed the existence of glioblastoma profile subgroups which correlated with prognostic factors. Moreover, we showed that the presence of circulating bevacizumab and carmustine did not alter this calorimetric signature of the disease, indicating that an evolution of the profile could be followed without being masked by ongoing systemic treatment. Thus, our results constitute a very promising proof of principle that a specific calorimetric profile could be detected in the plasma of glioblastoma patients. Moreover, we believe that our findings point to a potential easy-to-use non-invasive monitoring tool for glioblastoma patients., Competing Interests: CONFLICTS OF INTEREST The Authors declare no conflicts of interest.

Aggregation of TDP-43 (transactive response DNA binding protein 43 kDa) is a hallmark of certain forms of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Moreover, intracellular TDP-43-positive inclusions are often found in other neurodegenerative diseases. Recently it was shown that zinc ions can provoke the aggregation of endogenous TDP-43 in cells, allowing to assume a direct interaction of TDP-43 with zinc ions. In this work, we investigated zinc binding to the 102-269 TDP-43 fragment, which comprise the two RNA recognition motifs. Using isothermal titration calorimetry, mass spectrometry, and differential scanning fluorimetry, we showed that zinc binds to this TDP-43 domain with a dissociation constant in the micromolar range and modifies its tertiary structure leading to a decrease of its thermostability. Moreover, the study by dynamic light scattering and negative stain electron microscopy demonstrated that zinc ions induce auto-association process of this TDP-43 fragment into rope-like structures. These structures are thioflavin-T-positive allowing to hypothesize the direct implication of zinc ions in pathological aggregation of TDP-43.

Certain forms of amyotrophic lateral sclerosis (ALS) are associated with an altered compartmentalization of FUS and its aggregation in the cytoplasm of motoneurons. FUS is a DNA/RNA-binding protein that is involved in DNA repair and the regulation of transcription, splicing, RNA transport, and local translation. Two theories have been proposed to explain the mechanism of the pathophysiological process in ALS. The theories attribute degeneration of motor neurons to either loss or gain of FUS function. The review describes the main physiological functions of FUS and considers evidence for each of the theories of ALS pathogenesis.

α-Synuclein is involved in many important molecular processes in neuronal cells and their synapses, and its malfunction has been linked to the development of Parkinson's and certain other neurodegenerative diseases. Animal models allowing tightly monitored conditional inactivation of the encoding gene, Snca, are indispensible for studies aimed at understanding normal function of α-synuclein in various neuronal populations and its role in pathogenesis of neurodegenerative diseases. We have recently reported the production of several novel mouse lines for manipulating expression of the endogenous Snca gene, including a line for Cre-recombinase-driven conditional inactivation of the gene (mice with floxed Snca) and a new line with a constitutive knockout of α-synuclein. Rosa26-stop-lacZ reporter cassette is commonly used for monitoring efficiency of Cre-recombination but in mouse genome Snca and Rosa26 loci are located on the same chromosome. Here we describe production of lines with a modified Snca locus, either floxed or constitutively inactivated and the Rosa26-stop-lacZ reporter cassette located in cis on the mouse chromosome 6. These new mouse lines are invaluable for fast identification of cells with inactivation of Snca by Cre-recombination and represent useful tools for in vivo studies of α-synuclein function and dysfunction.

In this study, we analyzed serum for the presence of antibodies to gamma-synuclein in patients with amyotrophic lateral sclerosis (ALS) compared to the control group of patients with other neurological diseases and healthy control donors. As a result, antibodies against gamma-synuclein are not an ALS-specific feature and have been identified in patients with ALS as well as in the control group patients. Patients with the impaired cerebral circulation showed increased incidence of autoantibodies to gamma-synuclein, yet the difference lacks statistical representativeness due to limited sample size.

Biofilms of live bacteria forming on medical devices and implants contribute significantly to bacterial blood dissemination and to the spread of nosocomial infections. Cell surface SdrD protein plays a key role in the attachment of Staphylococcus aureus to the extracellular matrix (ECM) and in the formation of biofilm. SdrD binds calcium ions using its B1-B5 region bearing EF-hand Ca-binding sites, leading to conformational changes in the structure of SdrD. This alters the distance between the bacterial surface and the ECM-interacting domain of SdrD in a spring-like fashion, participating in bacterial attachment. In this study we investigated calcium binding to EF-hand sites of SdrD using isothermal titration calorimetry and determined the impact of this process on SdrD's thermodynamic stability. This allowed us to propose a model of B1-B5 reorganization upon binding of calcium and to get new insight into the molecular mechanism of SdrD's action.

Alpha-synuclein is a presynaptic protein of vertebrates that belongs to the family of synucleins. Normal functions of synucleins remain unknown. Alpha-synuclein is one of the causative factors of the familial and idiopathic forms of Parkinson's disease (PD). The progressive loss of dopaminergic (DA) neurons is characteristic of PD and the most severe damage occurs in the substantia nigra (SN). This leads to an erraticism of the synthesis and synaptic secretion of the neurotransmitters, subsequently resulting in the loss of the connections between brain areas. This work shows that alpha-synuclein is directly involved in the formation of the mature DA neurons of the midbrain at different stages of the ontogenesis and these findings are consistent with data obtained in other studies. Thus, alpha-synuclein may have a varying modulating effect on the growth dynamics and the fate of populations of DA neurons.