Reduced intranuclear mobility of APL fusion proteins accompanies their mislocalization and results in sequestration and decreased mobility of retinoid X receptor alpha.
Acute promyelocytic leukemia (APL) cells contain one of five chimeric retinoic acid alpha-receptor (RAR alpha) genes (X-RAR alpha) created by chromosomal translocations or deletion; each generates a fusion protein thought to transcriptionally repress RAR alpha target genes and block myeloid differentiation by an incompletely understood mechanism. To gain spatiotemporal insight into these oncogenic processes, we employed fluorescence microscopy and fluorescence recovery after photobleaching (FRAP). Fluorescence microscopy demonstrated that the intracellular localization of each of the X-RAR alpha proteins was distinct from that of RAR alpha and established which portion(s) of each X-RAR alpha protein-X, RAR, or both-contributed to its altered localization. Using FRAP, we demonstrated that the intranuclear mobility of each X-RAR alpha was reduced compared to that of RAR alpha. In addition, the mobility of each X-RAR alpha was reduced further by ligand addition, in contrast to RAR alpha, which showed no change in mobility when ligand was added. Both the reduced baseline mobility of X-RAR alpha and the ligand-induced slowing of X-RAR alpha could be attributed to the protein interaction domain contained within X. RXR alpha aberrantly colocalized within each X-RAR alpha; colocalization of RXR alpha with promyelocytic leukemia (PML)-RAR alpha resulted in reduced mobility of RXR alpha. Thus, X-RAR alpha may interfere with RAR alpha through its aberrant nuclear dynamics, resulting in spatial and temporal sequestration of RXR alpha and perhaps other nuclear receptor coregulators critical for myeloid differentiation.
Dong
Shuo
S
Section of Infectious Disease, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030, USA.
Stenoien
David L
DL
Qiu
Jihui
J
Mancini
Michael A
MA
Tweardy
David J
DJ
D021581:Active Transport, Cell Nucleus; D000818:Animals; D019556:COS Cells; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism*; D000071156:Promyelocytic Leukemia Protein; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism*; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins
D008024:Ligands; D008164:Luminescent Proteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion; D000071156:Promyelocytic Leukemia Protein; C000607071:RARA protein, human; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; D049452:Green Fluorescent Proteins