Differential roles of microtubule assembly and sliding in proplatelet formation by megakaryocytes

Megakaryocytes are terminally differentiated cells that, in their final hours, convert their cytoplasm into long, branched proplatelets, which remodel into blood platelets. Proplatelets elongate at an average rate of 0.85 μm/min in a microtubule-dependent process. Addition of rhodamine-tubulin to pe...

Full description

Saved in:
Bibliographic Details
Published in:Blood 2005-12, Vol.106 (13), p.4076-4085
Main Authors: Patel, Sunita R., Richardson, Jennifer L., Schulze, Harald, Kahle, Eden, Galjart, Niels, Drabek, Ksenija, Shivdasani, Ramesh A., Hartwig, John H., Italiano, Joseph E.
Format: Article
Language:English
Subjects:
Animals
Blood Platelets - cytology
Blood Platelets - metabolism
Cell Differentiation
Cytoplasm
Dynactin Complex
Dyneins - metabolism
Genes, Reporter - genetics
Hematopoiesis
Megakaryocytes - cytology
Megakaryocytes - metabolism
Mice
Microtubule-Associated Proteins - metabolism
Microtubules - metabolism
Protein Binding
Protein Subunits - metabolism
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Megakaryocytes are terminally differentiated cells that, in their final hours, convert their cytoplasm into long, branched proplatelets, which remodel into blood platelets. Proplatelets elongate at an average rate of 0.85 μm/min in a microtubule-dependent process. Addition of rhodamine-tubulin to permeabilized proplatelets, immunofluorescence microscopy of the microtubule plus-end marker end-binding protein 3 (EB3), and fluorescence time-lapse microscopy of EB3–green fluorescent protein (GFP)–expressing megakaryocytes reveal that microtubules, organized as bipolar arrays, continuously polymerize throughout the proplatelet. In immature megakaryocytes lacking proplatelets, microtubule plus-ends initiate and grow by centrosomal nucleation at rates of 8.9 to 12.3 μm/min. In contrast, plus-end growth rates of microtubules within proplatelets are highly variable (1.5-23.5 μm/min) and are both slower and faster than those seen in immature cells. Despite the continuous assembly of microtubules, proplatelets continue to elongate when net microtubule assembly is arrested. One alternative mechanism for force generation is microtubule sliding. Triton X-100–permeabilized proplatelets containing dynein and its regulatory complex, dynactin, but not kinesin, elongate with the addition of adenosine triphosphate (ATP) at a rate of 0.65 μm/min. Retroviral expression in megakaryocytes of dynamitin (p50), which disrupts dynactindynein function, inhibits proplatelet elongation. We conclude that while continuous polymerization of microtubules is necessary to support the enlarging proplatelet mass, the sliding of overlapping microtubules is a vital component of proplatelet elongation.
ISSN:0006-4971
1528-0020
DOI:10.1182/blood-2005-06-2204