A COMPUTER-ASSISTED SYSTEM FOR RECONSTRUCTING AND INTERPRETING THE DYNAMIC 3-DIMENSIONAL RELATIONSHIPS OF THE OUTER SURFACE, NUCLEUS AND PSEUDOPODS OF CRAWLING CELLS

Newly developed software additions to the three-dimensional dynamic im age analysis system, 3D-DIAS, are described for simultaneously reconst ructing and motion analyzing in three dimensions the outer surface, nu cleus and pseudopods of living, crawling cells. This new system is the n used to describe for the first time a nuclear behavior cycle in tran slocating Dictyostelium discoideum amoebae and to investigate the role of pseudopod extension in this process. The nuclear behavior cycle is tuned to the two phases of the general cell behavior cycle [Wessels e t al., 1994], and includes nuclear migration both in the z- and in the x,y-axes from the proximal border of the prior anterior pseudopod to the proximal border of a newly expanding anterior pseudopod. Nuclear m igration is cued by pseudopod-substratum contact, achieves velocities in excess of 50 mu m/min, and is accompanied by characteristic changes in nuclear shape. The rules and characteristics of nuclear behavior a re demonstrated to be intact in two mutants affecting pseudopod format ion, a myosin IB null mutant (myoB(-)) and a myosin II heavy chain pho sphorylation mutant (3XALA). The rules and characteristics of nuclear migration, however, are disrupted upon dissolution of microtubules by colcemid. Together the above results demonstrate that the newly develo ped 3D-DIAS system can be used to gain new insights into the dynamic c hanges in the intracellular 3D architecture associated with cellular t ranslocation. Cell Motil. Cytoskeleton 41:225-246, 1998. (C) 1998 Wile y-Liss, Inc.