Three-dimensional microscopy of the Rad51 recombination protein during meiotic prophase

An open question in meiosis is whether the Rad51 recombination protein func tions solely in meiotic recombination or whether it is also involved in the chromosome homology search. To address this question, we have performed th ree-dimensional high-resolution immunofluorescence microscopy to visualize native Rad51 structures in maize male meiocytes. Maize has two closely rela ted RAD51 genes that are expressed at low levels in differentiated tissues and at higher levels in mitotic and meiotic tissues. Cells and nuclei were specially fixed and embedded in polyacrylamide to maintain both native chro mosome structure and the three dimensionality of the specimens. Analysis of Rad51 in maize meiocytes revealed that when chromosomes condense during le ptotene, Rad51 is diffuse within the nucleus. Rad51 foci form on the chromo somes at the beginning of zygotene and rise to similar to 500 per nucleus b y mid-zygotene when chromosomes are pairing and synapsing. During chromosom e pairing, we consistently found two contiguous Rad51 fool on paired chromo somes. These paired foci may identify the sites where DNA sequence homology is being compared. During pachytene, the number of Rad51 foci drops to sev en to 22 per nucleus. This higher number corresponds approximately to the n umber of chiasmata in maize meiosis. These observations are consistent with a role for Rad51 in the homology search phase of chromosome pairing in add ition to its known role in meiotic recombination.