Use they are able to separate the two daughter Fenbutatin oxide Purity & Documentation nuclei solely by pulling forces exerted by way of astral microtubules, most like through minus-end directed motor activity of cortical dynein [237]. four. Centrosome-Nucleus Attachment Like all centrosomal structures in vegetative cells, the Dictyostelium centrosome is structurally linked to the cytosolic side on the nucleus during interphase. Not surprisingly, one particular key protein of this linkage will be the nuclear envelope protein Sun1, named right after the founding members from the Sun-family, i.e., fission yeast Sad1 and Caenorhabditis elegans UNC-84, which share a widespread Sun-domain. In most eukaryotes Sun1 is an inner nuclear membrane protein, forming a Quinacrine hydrochloride medchemexpress trimer and interacting, by means of its Sun-domain, using the so-called KASH-domain proteins (named right after Klarsicht, ANC-1, SYNE1 homology) within the perinuclear space [239]. Because the different KASH domain proteins interact directly or indirectly with all three cytoskeletal components (actin, microtubules, intermediate filaments) the term LINC complex (linker in the nucleus and cytoskeleton) was coined for the Sun/KASH domain protein heterodimer [240]. At the nuclear side, Sun1 interacts with lamins in animal cells as well as in Dictyostelium [241]. However, around the cytosolic face of the nuclear envelope the scenario in Dictyostelium appears to be unique. Sun1 is present in both nuclear membanes with no robust bias towards the inner nuclear membrane [124,125] and there isn’t any clear orthologue for a KASH domain protein. Because of its similarity to mammalian nesprins, the outer nuclear membrane protein interaptin was discussed as a Dictyostelium KASH domain protein [125,242]. But interaptin is undoubtedly no element of a LINC complicated, because it lacks the conserved KASH domain and definitely will not interact with Sun1 [125]. Sun1 is having said that required for centrosome/nucleus attachment. It co-purifies with isolated centrosomes and is concentrated at the nuclear envelope within the direct vicinity of your centrosome (Figure four). Sun1 mutants are defective in centrosome/nucleus attachment. It is feasible that the centrosome/nucleus linker employs Sun1 on both sides with the membrane, and that an unknown protein of the perinuclear space mediates this interaction. Even though a direct interaction with Sun1 remains to become proven, the unusual kinesin Kif9 is often a most likely candidate to get a LINC complicated element in Dictyostelium. Kif9 is definitely an internal motor kinesin, which could be grouped into the kinesin-13 family members, which ordinarily act as microtubule depolymerases [130]. Within this group Kif9 is distinctive in containing a 23 residue transmembrane domain close to its C-terminal finish, targeting the protein towards the outer nuclear envelope exactly where it accumulates within the pericentrosomal area. Knockout of Kif9 disrupts the centrosome/nucleus linkage and causes dispersal of Sun1, away from the pericentrosomal region with the nuclear envelope [130].Figure 4. Centrosome-Nucleus-Centromere cluster. (A) Immunoelectron microscopy image displaying one particular section of an isolated nucleus using the attached centrosome. Nuclei had been labeled with an antibody against Dictyostelium Sun1 and nanogold conjugated anti-rabbit antibodies. The centrosome (Cn), the centromeric cluster (Cm), the nuclear envelope (NE) plus the endoplasmic reticulum (ER) are indicated (image by Prof. Otto Baumann); (B) Immunofluorescence microscopy image of a Sun1-GFP knock-in cell (green) stained with an antibody against the centrosomal core protein CP91 and anti-rabbit-AlexaFluor 568 conjug.
