RESUMEN
Based on the protein kinase A (PKA)/GSK3ß interaction protein (GSKIP)/glycogen synthase kinase 3ß (GSK3ß) axis, we hypothesized that these might play a role in Tau phosphorylation. Here, we report that the phosphorylation of Tau Ser409 in SHSY5Y cells was increased by overexpression of GSKIP WT more than by PKA- and GSK3ß-binding defective mutants (V41/L45 and L130, respectively). We conducted in vitro assays of various kinase combinations to show that a combination of GSK3ß with PKA but not Ca2+/calmodulin-dependent protein kinase II (CaMK II) might provide a conformational shelter to harbor Tau Ser409. Cerebrospinal fluid (CSF) was evaluated to extend the clinical significance of Tau phosphorylation status in Alzheimer's disease (AD), neurological disorders (NAD), and mild cognitive impairment (MCI). We found higher levels of different PKA-Tau phosphorylation sites (Ser214, Ser262, and Ser409) in AD than in NAD, MCI, and normal groups. Moreover, we used the CRISPR/Cas9 system to produce amyloid precursor protein (APPWT/D678H) isogenic mutants. These results demonstrated an enhanced level of phosphorylation by PKA but not by the control. This study is the first to demonstrate a transient increase in phosphor-Tau caused by PKA, but not GSK3ß, in the CSF and induced pluripotent stem cells (iPSCs) of AD, implying that both GSKIP and GSK3ß function as anchoring proteins to strengthen the cAMP/PKA/Tau axis signaling during AD pathogenesis.
RESUMEN
GSK3ß interacting protein (GSKIP) is a naturally occurring negative regulator of GSK3ß and retains both the Protein Kinase A Regulatory subunit binding (PKA-RII) domain and GSK3ß interacting domain. Of these two domains, we found that PKA-RII is required for forming a working complex comprising PKA/GSKIP/GSK3ß/Drp1 to influence phosphorylation of Drp1 Ser637. In this study, bioinformatics and experimental explorations re-analyzing GSKIP's biofunctions suggest that the evolutionarily conserved Domain of Unknown Function (DUF727) is an ancestral prototype of GSKIP in prokaryotes, and acquired the C-terminal GSK3ß binding site (tail) in invertebrates except for Saccharomyces spp., after which the N-terminal PKA-RII binding region (head) evolved in vertebrates. These two regions mutually influence each other and modulate GSKIP binding to GSK3ß in yeast two-hybrid assays and co-immunoprecipitation. Molecular modeling showed that mammalian GSKIP could form a dimer through the L130 residue (GSK3ß binding site) rather than V41/L45 residues. In contrast, V41/L45P mutant facilitated a gain-of-function effect on GSKIP dimerization, further influencing binding behavior to GSK3ß compared to GSKIP wild-type (wt). The V41/L45 residues are not only responsible for PKA RII binding that controls GSK3ß activity, but also affect dimerization of GSKIP monomer, with net results of gain-of-function in GSKIP-GSK3ß interaction. In addition to its reported role in modulating Drp1, Ser637 phosphorylation caused mitochondrial elongation; we postulated that GSKIP might be involved in the Wnt signaling pathway as a scavenger to recruit GSK3ß away from the ß-catenin destruction complex and as a competitor to compete for GSK3ß binding, resulting in accumulation of S675 phosphorylated ß-catenin.
Asunto(s)
Proteínas Represoras/química , Proteínas Represoras/metabolismo , Vía de Señalización Wnt , Sitios de Unión , Biología Computacional , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Dinaminas , Evolución Molecular , GTP Fosfohidrolasas/química , GTP Fosfohidrolasas/metabolismo , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Células HEK293 , Humanos , Proteínas Asociadas a Microtúbulos/química , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Modelos Moleculares , Fosforilación , Filogenia , Unión Proteica , Dominios Proteicos , Multimerización de Proteína , Proteínas Represoras/genética , Serina/química , Técnicas del Sistema de Dos HíbridosRESUMEN
INTRODUCTION: In Taiwan, In Vitro Diagnostic Medical Device (IVD) is regulated as medical device since 1987, and the implementation of IVD registration was fully completed in 2005. The management system of IVD medical device is highly similar with a guidance 'The GHTF Regulatory Model' developed by Global Harmonization Task Force (GHTF) in 2011 for use of regulation development on medical devices. Area covered: In this study, the Regulatory Model developed by GHTF was compared with Taiwanese IVD management system and it has shown that these two regulatory frameworks are highly similar. Expert commentary: The experience of IVD management in Taiwan can serve a strong evidence to prove the feasibility and effectiveness of GHTF Regulatory Model.
Asunto(s)
Aprobación de Recursos , Juego de Reactivos para Diagnóstico , Humanos , Modelos Teóricos , TaiwánRESUMEN
Group B streptococcus (GBS) is a major cause of severe systemic infections among the newborn. Both recurrent and maternal mastitis-associated, group B streptococcus diseases are uncommon. Persistence of GBS colonization of infants' mucous membrane is postulated to influence the pathogeneses of recurrent GBS infection. The authors describe a term infant who was treated for GBS sepsis and meningitis and then later developed recurrent GBS sepsis, without meningitis, due to feeding of infected breast milk. Randomly amplified polymorphic DNA polymerase chain reaction assay was performed to demonstrate that the GBS isolates from the first and second episode of infection and the maternal milk are identical. The authors conclude that transmission of GBS through breast milk should be considered in cases of recurrent neonatal GBS infection and bacterial culture of breast milk should be routinely performed in such cases.
Asunto(s)
Lactancia Materna , Transmisión Vertical de Enfermedad Infecciosa , Infecciones Estreptocócicas/transmisión , Streptococcus agalactiae , Ampicilina/uso terapéutico , Antibacterianos/uso terapéutico , Preescolar , Femenino , Estudios de Seguimiento , Humanos , Leche Humana/microbiología , Reacción en Cadena de la Polimerasa , Recurrencia , Infecciones Estreptocócicas/tratamiento farmacológicoAsunto(s)
Neoplasias Óseas/diagnóstico , Neoplasias Óseas/genética , Reordenamiento Génico , Linfoma no Hodgkin/diagnóstico , Linfoma no Hodgkin/genética , VDJ Recombinasas/genética , Adulto , Secuencia de Bases , Progresión de la Enfermedad , Genes de Inmunoglobulinas , Humanos , Masculino , Datos de Secuencia Molecular , Metástasis de la Neoplasia , Reacción en Cadena de la Polimerasa , RecurrenciaRESUMEN
BACKGROUND: The clinicopathologic and molecular features of follicular lymphoma (FL) in Taiwan have not been well defined. We conducted a retrospective study including history review, immunohistochemistry, and molecular study for the major breakpoint region (MBR) of t(14;18) and correlated these findings with survival. PATIENTS AND METHODS: Sixty-five FLs were identified, with a male to female ratio of 1.9:1 and a median age of 63 years (mean, 60 years). Sixty cases (92%) were nodal, 4 (6%) were extranodal, and 1 (2%) was indeterminate. The median ages of the nodal and extranodal cases were 63 years and 44 years, respectively. Disease staging in 59 patients included 15 patients (25%) with stage I disease, 14 (24%) with stage II, 20 (34%) with stage III, and 10 (17%) with stage IV. Forty-four patients received chemotherapy, 2 patients received chemotherapy with palliative radiation therapy, and 13 patients received supportive treatment/observation. RESULTS: The 5-year survival rate was 52.6%. The cases were classified as grades 1 (n = 27; 42%), 2 (n = 22; 34%), 3A (n = 13; 20%), and 3B (n = 3; 5%). Twenty cases (31%) were positive for MBR, including 19 of 57 (33%) nodal cases and 1 of 4 (25%) primary extanodal FLs. Patients with low-stage disease (stages I/II) had a better survival rate than patients with high-stage disease (III/IV; log-rank test, P = 0.012). CONCLUSION: This is the largest series of Taiwanese FLs with immunophenotypes and MBR detection rates similar to those of the West. Disease stage was statistically significant with regard to survival. Although the number of extranodal FLs cases was small, the patients were younger, their tumors had lower CD10 expression, and they had more favorable survival rates than patients with nodal disease.