Summary Lamins A, C, B1 and B2 form nuclear intermediate filaments as major components of the dynamic genome- associated nucleoskeleton. Lamins associate with nuclear envelope (NE) membrane proteins, together forming nuclear ‘lamina’ networks. Lamins and key partners (LEM-domain proteins and BANF1) are essential during exit from mitosis to ensure that chromosomes are coalesced, captured and properly organized within the daughter nucleus. During interphase, nuclear lamina networks have fascinating roles in the higher-order architecture of transcriptionally-inactive regions of the genome (heterochromatin). Silent regions of each chromosome, known as Lamina Associated Domains (LADs), are typically located near the NE. There are clear correlations between LAD organization, epigenomic regulation, and the functional three-dimensional (3D) folding of the genome. A-type lamins (encoded by LMNA) have key roles in LAD organization. LMNA gives rise to two major somatic isoforms, lamin A and lamin C, by alternative mRNA splicing. Because the first 566 residues of human lamin A and lamin C are identical, they were long thought to function redundantly. However new reports show lamin A and lamin C form separate filaments, associate differentially with nuclear pore complexes and have distinct metabolic phenotypes. We discovered lamin C is required for LADs to associate with the NE during interphase. Furthermore, lamin C is specifically and strikingly nucleoplasmic during telophase and early-G1, in stark contrast to lamin A at the nascent NE. Although lamin C is not LAD- associated in early-G1, we found lamin C associates with LADs as they return to their ‘tethered’ positions at the NE. We propose lamin C is required for LAD recruitment to the NE, and will test this hypothesis in cells specifically downregulated for lamin C or lamin A. We can detect distinct yet overlapping proteomes in unsynchronized cells, comprising emerin and LAP2beta at the nuclear membrane, lamins and soluble partners (‘connectome’), and a novel LAD-associated proteome. We hypothesize that lamin C specifically interacts with LADs or LAD-associated proteins during exit from mitosis as a pathway to re-establish the tissue-specific positioning of silent chromatin (LADs) at the NE. Our models predict distinct proteomes for lamin C vs lamin A during mitotic exit, distinct changes in the LAD proteome during mitotic exit, and perturbed LAD organization or LAD recruitment to the NE in cells that lack lamin C during mitotic exit. We will test these models by super- resolution imaging of lamins and LADs in single cells, directed proteomics, genome organization mapping and functional studies in cells downregulated for either lamin C, lamin A or validated proteins identified in this work. This work is expected to fill major gaps in understanding how genome architecture is established after mitosis, and functional differences between lamin A and lamin C that may also be relevant to the mechanisms of dis...