Related Experiment Videos
Identification of PML oncogenic domains (PODs) in human megakaryocytes
A Drouin1, A Schmitt, J M Massé
1Institut Cochin de Génétique Moléculaire, INSERM U. 474, Paris, France.
Abstract:
Megakaryocytes (Mks) are unique cells in the human body in that they carry a single and polyploid nucleus. It is therefore of interest to understand their nuclear ultrastructure. PML oncogenic domains (PODs) were described in several types of eukaryotic cells using human autoantibodies which recognize nuclear antigens with a specific speckled pattern (dots) in indirect immunofluorescence (IF). Two main antigens, PML and Sp 100, usually colocalize and concentrate in these nuclear subdomains. We investigated the presence of PODs using IF and immunoelectron microscopy (IEM) in cells from megakaryocytic lineage: the HEL cell line and human cultured Mks. Antibodies against PML, Sp100, and anti-nuclear dots were used in single and double labeling. PODs were identified in HEL cells and in human Mks, and their ultrastructure was characterized. We then used IF to quantify PODs within Mks and showed that their number increased proportionally to nuclear lobularity. In summary, we report the identification of PODs in human Mks at an ultrastructural level and an increase in PODs number in parallel with Mk ploidy. We show that endomitosis not only leads to DNA increase but also to the multiplication of at least one of the associated nuclear structures.
Insights
PML oncogenic domains (PODs) were identified in human megakaryocytes (Mks). Their numbers increase with nuclear lobularity and ploidy during megakaryocyte development.
Area of Science:
- Cell Biology
- Nuclear Structure
- Hematopoiesis
Background:
- Megakaryocytes (Mks) possess unique polyploid nuclei.
- PML oncogenic domains (PODs) are nuclear subdomains found in various eukaryotic cells.
- PODs are typically characterized by PML and Sp 100 antigens, appearing as speckled dots.
Purpose of the Study:
- To investigate the presence and ultrastructure of PODs in megakaryocytic lineage cells.
- To determine the relationship between PODs and megakaryocyte nuclear morphology, specifically ploidy and lobularity.
Main Methods:
- Indirect immunofluorescence (IF) and immunoelectron microscopy (IEM) were employed.
- Antibodies against PML, Sp 100, and anti-nuclear dots were used for labeling.
- HEL cell line and human cultured Mks were utilized for the study.
Main Results:
- PODs were successfully identified in both HEL cells and human Mks at the ultrastructural level.
- The number of PODs was found to increase proportionally with nuclear lobularity in Mks.
- A parallel increase in PODs number was observed with megakaryocyte ploidy.
Conclusions:
- This study reports the ultrastructural identification of PODs in human megakaryocytes.
- Megakaryocyte endomitosis leads to not only DNA increase but also the multiplication of nuclear structures like PODs.
- PODs are dynamic nuclear components that expand with megakaryocyte ploidy and nuclear complexity.