At the physiological level, PML has been functionally linked to anti-inflammatory and antiviral response pathways, metabolism, stem cell maintenance, and aging, while more mechanistically, PMLвЂ™s role in tumor suppression is linked to control of the cell cycle, apoptosis/senescence, cell migration, angiogensis, and the DNA damage response ( 9, 10). Therefore, PML is regarded as a potent tumor suppressor in in vitro (biochemistry, cell culture experiments) and in vivo (model organisms). Furthermore, loss of PML is a hallmark of human cancers from diverse tissues ( 8). PML knock-out mice develop a range of cancers including papillomas, carcinomas, and lymphomas after exposure to carcinogens ( 7). Overexpression of PML in cancer cell lines induces cell cycle arrest and apoptosis ( 6). PML NBs are heterogeneous and dynamic structures, ranging in size from 0.1 to 1.0 Вµm, and typically there are 5вЂ“30 bodies per nucleus, depending on the cell type, phase of cell cycle, and the cellular stress level ( 3вЂ“ 5). PML NBs appear as nuclear dot-shaped structures that are interspersed between chromatin ( 2). PML protein is the major building unit of the so-called PML nuclear bodies (PML NBs). The promyelocytic leukemia ( pml) gene is a target of the t(15 17) chromosomal translocation, which fuses pml reciprocally with retinoic acid receptor О± (RARО±) ( 1). Since most if not all of the microscopic equipment to perform these techniques may be available in an institutional or nearby facility, we hope to encourage more researches to exploit sophisticated imaging tools for their research in cancer biology. These include fluorescence redistribution after photobleaching, fluorescence resonance energy transfer, fluorescence correlation spectroscopy, raster image correlation spectroscopy, ultraviolet laser microbeam-induced DNA damage, erythrocyte-mediated force application, and super-resolution microscopy approaches. In this report, we summarize the current knowledge on PML nuclear bodies and describe several fluorescence imaging, manipulation, FFM, and super-resolution techniques suitable to analyze PML body assembly and function. In parallel, a variety of fluorescence fluctuation microscopy (FFM) approaches have been developed which allow precise determination of the biophysical and interaction properties of cellular factors at the single molecule level in living cells. With the advent of genetically encoded fluorescent proteins it became possible to trace protein function in living specimens.

It remains a big challenge to unify all the different previously suggested cellular functions of PML NBs into one mechanistic model. However, despite extensive research over the past decades, the precise biochemical function of PML in these pathways is still elusive. While PML NBs have been implicated in many cellular functions including cell cycle regulation, survival and apoptosis their role as signaling hubs along major genome maintenance pathways emerged more clearly. In the cell nucleus, PML seeds the formation of macromolecular multiprotein complexes, known as PML nuclear bodies (PML NBs). The promyelocytic leukemia ( pml) gene product PML is a tumor suppressor localized mainly in the nucleus of mammalian cells.

2Carl Zeiss Microscopy GmbH, Jena, Germany.1Leibniz Institute on Aging Research, Jena, Germany.

Carl zeiss microscope models structural