![]() ![]() However, elucidating the mechanism of MTOC assembly is important since MTOCs perform a variety of functions in the cell, including cell signaling, cilia assembly, intraflagellar transport, and chromosome segregation. Few methods are available to examine large protein structures such as microtubule-organizing centers (MTOCs) in intact cells. The goal of molecular, biochemical and cell biological studies is to elucidate the function of cellular components and understand how proteins and the complexes they form interact in vivo. makes it possible to carry out studies that examine the movements of whole protein complexes during cell division. Until now, highly detailed surveys of protein structures have been limited to a handful of proteins and conditions. The next challenges are to understand the details of how this works and to use the same method to study other large protein complexes in cells. observed that this set of proteins interact with the new spindle pole as it forms, instead of afterwards as was previously believed.īurns et al.'s findings suggest that the spindle pole body assembles into the membrane surrounding the nucleus at the same time as it is copied. Some of these proteins enable the spindle pole to insert into the membrane that surrounds the cell's nucleus. The experiments mapped the positions of 18 proteins within the spindle pole body during its duplication. developed a new method to study the spindle pole body using fluorescent protein tags and a sophisticated microscopy technique. The spindle pole in yeast-known as the spindle pole body-is an ideal model to study this problem because the proteins that form it have already been identified and it is easy to study yeast in the laboratory.īurns et al. While we understand how DNA is copied, we do not know how cells copy proteins. To ensure that the DNA is separated into two equal parts, the microtubules must emerge from two points in the cell, known as spindle poles.Įach spindle pole is made of a group (or ‘complex’) of proteins and these have to be copied before the cell can divide. First, the DNA of the parent cell is copied, then it must be physically separated into the daughter cells by a structure made of filaments called microtubules. Our observation that proteins involved in membrane insertion, such as Mps2, Bbp1, and Ndc1, also accumulate at the new SPB early in duplication suggests that SPB assembly and NE insertion are coupled events during SPB formation in wild-type cells.Ĭells divide to produce two new daughter cells that each contain the same genetic material. The increased resolution and quantitative intensity information obtained using this method allowed us to demonstrate that SPB duplication begins by formation of an asymmetric Sfi1 filament at mitotic exit followed by Mps1-dependent assembly of a Spc29- and Spc42-dependent complex at its tip. To better understand this process, we developed a novel two-color structured illumination microscopy with single-particle averaging (SPA-SIM) approach to study the localization of all 18 SPB components during duplication using endogenously expressed fluorescent protein derivatives. ![]() Duplication of the yeast centrosome (called the spindle pole body, SPB) is thought to occur through a series of discrete steps that culminate in insertion of the new SPB into the nuclear envelope (NE). ![]()
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