Use they are able to separate the two daughter nuclei solely by pulling forces exerted by means of astral microtubules, most like by means of 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 towards the cytosolic side of your nucleus for the duration of interphase. Not surprisingly, one particular key protein of this linkage is the nuclear envelope protein Sun1, named just after the founding members in the Sun-family, i.e., fission yeast Sad1 and Caenorhabditis elegans UNC-84, which share a popular Sun-domain. In most eukaryotes Sun1 is definitely an inner nuclear membrane protein, forming a trimer and interacting, via its Sun-domain, with the so-called KASH-domain proteins (named just after Klarsicht, ANC-1, SYNE1 homology) within the perinuclear space [239]. Since the many KASH domain proteins interact directly or indirectly with all three cytoskeletal elements (actin, microtubules, intermediate filaments) the term LINC complex (linker of your nucleus and cytoskeleton) was coined for the Sun/KASH domain protein heterodimer [240]. In the nuclear side, Sun1 interacts with lamins in animal cells and also in Dictyostelium [241]. However, around the cytosolic face of the nuclear envelope the RHPS4 MedChemExpress circumstance in Dictyostelium seems to become one of a kind. Sun1 is present in both nuclear membanes with no strong bias towards the inner nuclear membrane [124,125] and there is no clear orthologue for any 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 definitely no portion of a LINC complex, because it lacks the conserved KASH domain and definitely doesn’t interact with Sun1 [125]. Sun1 is having said that expected for centrosome/nucleus attachment. It co-purifies with isolated centrosomes and is concentrated at the nuclear envelope within the direct vicinity on the centrosome (Figure four). Sun1 mutants are defective in centrosome/nucleus attachment. It is actually probable 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. 5′-O-DMT-2′-O-TBDMS-Bz-rC In Vivo Though a direct interaction with Sun1 remains to become established, the uncommon kinesin Kif9 is really a most likely candidate to get a LINC complex component in Dictyostelium. Kif9 is an internal motor kinesin, which can be grouped into the kinesin-13 loved ones, which ordinarily act as microtubule depolymerases [130]. Inside this group Kif9 is distinctive in containing a 23 residue transmembrane domain close to its C-terminal end, targeting the protein towards the outer nuclear envelope where it accumulates inside the pericentrosomal region. Knockout of Kif9 disrupts the centrosome/nucleus linkage and causes dispersal of Sun1, away in the pericentrosomal area from the nuclear envelope [130].Figure 4. Centrosome-Nucleus-Centromere cluster. (A) Immunoelectron microscopy image displaying one 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) along with 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.