ABSTRACT
Chromosome segregation requires assembly of kinetochores on centromeric chromatin to mediate interactions with spindle microtubules and control cell-cycle progression. To elucidate the protein architecture of human kinetochores, we developed a two-color fluorescence light microscopy method that measures average label separation, Delta, at <5 nm accuracy. Delta analysis of 16 proteins representing core structural complexes spanning the centromeric chromatin-microtubule interface, when correlated with mechanical states of spindle-attached kinetochores, provided a nanometer-scale map of protein position and mechanical properties of protein linkages. Treatment with taxol, which suppresses microtubule dynamics and activates the spindle checkpoint, revealed a specific switch in kinetochore architecture. Cumulatively, Delta analysis revealed that compliant linkages are restricted to the proximity of chromatin, suggested a model for how the KMN (KNL1/Mis12 complex/Ndc80 complex) network provides microtubule attachment and generates pulling forces from depolymerization, and identified an intrakinetochore molecular switch that may function in controlling checkpoint activity.
Subject(s)
Kinetochores/chemistry , Kinetochores/metabolism , Microtubules/chemistry , Microtubules/metabolism , Cytoskeletal Proteins , DNA-Binding Proteins/metabolism , HeLa Cells , Humans , Metaphase , Microscopy, Fluorescence , Microtubule-Associated Proteins/metabolism , Nuclear ProteinsABSTRACT
BACKGROUND: Psoriasis is being recognized as an autoimmune disease in which immunocyte-derived cytokines are thought to drive the development of the altered keratinocyte phenotype. Although the role of tumor necrosis factor alpha (TNF-alpha) in psoriasis is not completely understood, it may underlie many of the key steps that lead to induction and maintenance of the disease. Infliximab is an immunoglobulin monoclonal antibody that binds and inactivates TNF-alpha and has been successfully used in the management of TNF-alpha-mediated diseases, such as Crohn disease and rheumatoid arthritis. OBSERVATIONS: Two patients with recalcitrant psoriasis that was unresponsive to multiple skin-directed and systemic therapies were treated with a single infusion of infliximab. The treatments resulted in rapid and complete clearing of psoriatic erythroderma and resolution of symptoms of arthritis in one case and complete clearing of widespread psoriatic plaques and improvement of symptoms of arthritis and inflammatory bowel disease in the other. The single treatments with infliximab were well tolerated with no immediate or long-term adverse effects noted. CONCLUSION: A single infusion of infliximab at 5 to 10 mg/kg resulted in the rapid and complete clearing of recalcitrant psoriatic plaques and erythroderma with a disease-free interval of 3 to 4 months in these 2 patients and improved the symptoms of psoriatic arthritis.
