Proteasome-mediated degradation of integral inner nuclear membrane protein emerin in fibroblasts lacking A-type lamins

Abstract

We previously identified and characterized a homozygous LMNA nonsense mutation leading to the absence of A-type lamins in a premature neonate who died at birth. We show here that the absence of A-type lamins is due to degradation of the aberrant mRNA transcript with a premature termination codon. In cultured fibroblasts from the subject with the homozygous LMNA nonsense mutation, there was a decreased steady-state expression of the integral inner nuclear membrane proteins emerin and nesprin-1α associated with their mislocalization to the bulk endoplasmic reticulum and a hyperphosphorylation of emerin. To determine if decreased emerin expression occurred posttranslationally, we treated cells with a selective proteasome inhibitor and observed an increase in expression. Our results show that mislocalization of integral inner nuclear membrane proteins to the endoplasmic reticulum in human cells lacking A-type lamins leads to their degradation and provides the first evidence that their degradation is mediated by the proteasome. KeywordsNuclear envelope; Lamin; Emerin; Nesprin; Proteasome; Muscular dystrophy Lamins are intermediate filament proteins that form the nuclear lamina, a fibrous meshwork underlying the inner nuclear membrane of higher eukaryotic cells [1]. In human, two types of lamins are expressed in somatic cells according to their sequences and biochemical properties, A-type and B-type. Lamins A and C, encoded by the LMNA gene, are the predominant A-type lamins in mammalian somatic cells and arise from alternative RNA splicing. Two genes, LMNB1 and LMNB2, respectively, encode the somatic mammalian B-type lamins B1 and B2 [1]. While lamins B1 and B2 are expressed in most of all somatic cells, the expression of Atype lamins is developmentally regulated and generally restricted to differentiated cells [2,3]. The nuclear lamina is attached to the inner nuclear membrane via interactions with integral proteins, among them emerin, nesprin-1α, LBR, and LAPs [4]. Lamins A and C also interact

Functional Characterization of a Novel Truncating Mutation in Lamin A/C Gene in a Family with a Severe Cardiomyopathy with Conduction Defects

“…Of note, the expression of mCh-D243Gfs*4 into the ER did not alter the localization of other nuclear envelope proteins such as emerin (Fig. 3B), as showed for other truncating LMNA mutations [20]. Since both the D243Gfs*4 and the previously studied R321X [8] share the same peculiar ER localization, we supposed for these truncating mutations common pathogenic mechanisms once transfected in HEK293 cells.…”

LULL1 Retargets TorsinA to the Nuclear Envelope Revealing an Activity That Is Impaired by theDYT1Dystonia Mutation

“…Western blot analysis of Sun2 in such cells confirms a decrease in the overall level of Sun2 but not nucleoporins or a general ER marker (Figure 7). Increased degradation of INM proteins has been reported when lamin anchors are absent (Muchir et al, 2006); it is possible that TorA could similarly destabilize Sun2 as a consequence of altering its association with the nuclear lamina.…”

Lamins, laminopathies and disease mechanisms: Possible role for proteasomal degradation of key regulatory proteins

“…In fibroblasts from an LGMD1B patient with a homozygous LMNA nonsense mutation (Y259X), which leads to absence of lamin A, the integral membrane proteins emerin and nesprin-1α are mislocalized to the ER and subsequently degraded; this degradation is mediated by the proteasomal machinery (Muchir et al 2006). Proteomics analysis has demonstrated that reduction of lamin A/C to~10% of normal values by an shRNA approach in HeLa cells leads to depletion of 34 proteins, most of which are involved in cytoskeletal organization, cell cycle regulation and proliferation (Chen et al 2009).…”