Lamin: Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References

Gene name - Lamin

Synonyms - Lamin Dm0

Cytological map position - 25F1--25F2

Function - intermediate filament, chromatin associated protein

Keywords - cytoskeleton, chromatin associated proteins

Symbol - Lam

FlyBase ID: FBgn0002525

Genetic map position - 2-[17].

Classification - nuclear lamin

Cellular location - nuclear



NCBI links: Precomputed BLAST | Entrez Gene

Recent literature
Chen, H., Zheng, X. and Zheng, Y. (2015). Lamin-B in systemic inflammation, tissue homeostasis, and aging. Nucleus [Epub ahead of print]. PubMed ID: 25875575
Summary:
Gradual loss of tissue function (or homeostasis) is a natural process of aging and is believed to cause many age-associated diseases. In human epidemiology studies, the low-grade and chronic systemic inflammation in elderly has been correlated with the development of aging related pathologies. Although it is suspected that tissue decline is related to systemic inflammation, the cause and consequence of these aging phenomena are poorly understood. By studying the Drosophila fat body and gut, this study has uncovered a mechanism by which lamin-B loss in the fat body upon aging induces age-associated systemic inflammation. This chronic inflammation results in the repression of gut local immune response, which in turn leads to the over-proliferation and mis-differentiation of the intestinal stem cells, thereby resulting in gut hyperplasia. Implications and remaining questions are discussed in light of these new observations.

Uchino, R., Sugiyama, S., Katagiri, M., Chuman, Y. and Furukawa, K. (2016). Non-farnesylated B-type lamin can tether chromatin inside the nucleus and its chromatin interaction requires the Ig-fold region. Chromosoma [Epub ahead of print]. PubMed ID: 26892013
Summary:
Lamins are thought to direct heterochromatin to the nuclear lamina (NL); however, this function of lamin has not been clearly demonstrated in vivo. To address this, polytene chromosome morphology were analyzed when artificial lamin variants were expressed in Drosophila endoreplicating cells. The CaaX-motif-deleted B-type lamin Dm0, but not A-type lamin C, was able to form a nuclear envelope-independent layer that was closely associated with chromatin. Other nuclear envelope proteins were not detected in this "ectopic lamina," and the associated chromatin showed a repressive histone modification marker but not a permissive histone modification marker nor RNA polymerase II proteins. Furthermore, deletion of the C-terminal lamin-Ig-fold domain prevents chromatin association with this ectopic lamina. Thus, non-farnesylated B-type lamin Dm0 can form an ectopic lamina and induce changes to chromatin structure and status inside the interphase nucleus.

Chen, H., Zheng, X., Xiao, D. and Zheng, Y. (2016). Age-associated de-repression of retrotransposons in the Drosophila fat body, its potential cause and consequence. Aging Cell [Epub ahead of print]. PubMed ID: 27072046
Summary:
Eukaryotic genomes contain transposable elements (TE) that can move into new locations upon activation. Since uncontrolled transposition of TEs, including the retrotransposons and DNA transposons, can lead to DNA breaks and genomic instability, multiple mechanisms, including heterochromatin-mediated repression, have evolved to repress TE activation. Studies in model organisms have shown that TEs become activated upon aging as a result of age-associated deregulation of heterochromatin. Considering that different organisms or cell types may undergo distinct heterochromatin changes upon aging, it is important to identify pathways that lead to TE activation in specific tissues and cell types. Through deep sequencing of isolated RNAs, this study reports an increased expression of many retrotransposons in the old Drosophila fat body, an organ equivalent to the mammalian liver and adipose tissue. This de-repression correlates with an increased number of DNA damage foci and decreased level of Drosophila lamin-B in the old fat body cells. Depletion of the Drosophila lamin-B in the young or larval fat body results in a reduction of heterochromatin and a corresponding increase in retrotransposon expression and DNA damage. Further manipulations of lamin-B and retrotransposon expression suggest a role of the nuclear lamina in maintaining the genome integrity of the Drosophila fat body by repressing retrotransposons.

Fabbretti, F., Iannetti, I., Guglielmi, L., Perconti, S., Evangelistella, C., Proietti De Santis, L., Bongiorni, S. and Prantera, G. (2016). Confocal analysis of nuclear lamina behavior during male meiosis and spermatogenesis in Drosophila melanogaster. PLoS One 11: e0151231. PubMed ID: 26963718
Summary:
Lamin family proteins are structural components of a filamentous framework, the nuclear lamina (NL), underlying the inner membrane of nuclear envelope. The NL not only plays a role in nucleus mechanical support and nuclear shaping, but is also involved in many cellular processes including DNA replication, gene expression and chromatin positioning. Spermatogenesis is a very complex differentiation process in which each stage is characterized by nuclear architecture dramatic changes, from the early mitotic stage to the sperm differentiation final stage. Nevertheless, very few data are present in the literature on the NL behavior during this process. This study shows the first and complete description of NL behavior during meiosis and spermatogenesis in Drosophila melanogaster. By confocal imaging, the NL modifications wee characterized from mitotic stages, through meiotic divisions to sperm differentiation with an anti-laminDm0 antibody against the major component of the Drosophila NL. It was observed that continuous changes in the NL structure occurred in parallel with chromatin reorganization throughout the whole process and that meiotic divisions occurred in a closed context. Finally, NL was examined in solofuso meiotic mutant, where chromatin segregation is severely affected, and a strict correlation was found between the presence of chromatin and that of NL.

Chen, H., Zheng, X., Xiao, D. and Zheng, Y. (2016). Age-associated de-repression of retrotransposons in the Drosophila fat body, its potential cause and consequence. Aging Cell [Epub ahead of print]. PubMed ID: 27072046
Summary:
Eukaryotic genomes contain transposable elements (TE) that can move into new locations upon activation. Since uncontrolled transposition of TEs, including the retrotransposons and DNA transposons, can lead to DNA breaks and genomic instability, multiple mechanisms, including heterochromatin-mediated repression, have evolved to repress TE activation. Studies in model organisms have shown that TEs become activated upon aging as a result of age-associated deregulation of heterochromatin. Considering that different organisms or cell types may undergo distinct heterochromatin changes upon aging, it is important to identify pathways that lead to TE activation in specific tissues and cell types. Through deep sequencing of isolated RNAs, this study report an increased expression of many retrotransposons in the old Drosophila fat body, an organ equivalent to the mammalian liver and adipose tissue. This de-repression correlates with an increased number of DNA damage foci and decreased level of Drosophila lamin-B in the old fat body cells. Depletion of the Drosophila lamin-B in the young or larval fat body results in a reduction of heterochromatin and a corresponding increase in retrotransposon expression and DNA damage. Further manipulations of lamin-B and retrotransposon expression suggest a role of the nuclear lamina in maintaining the genome integrity of the Drosophila fat body by repressing retrotransposons.

Hayashi, D., Tanabe, K., Katsube, H. and Inoue, Y. H. (2016). B-type nuclear lamin and the nuclear pore complex Nup107-160 influences maintenance of the spindle envelope required for cytokinesis in Drosophila male meiosis. Biol Open [Epub ahead of print]. PubMed ID: 27402967
Summary:
In higher eukaryotes, nuclear envelope (NE) disassembly allows chromatin to condense and spindle microtubules to access kinetochores. The nuclear lamina, which strengthens the NE, is composed of a polymer meshwork made of A- and B-type lamins. This study found that the B-type lamin (Lam) is not fully disassembled and continues to localize along the spindle envelope structure during Drosophila male meiosis I, while the A-type lamin (LamC) is completely dispersed throughout the cytoplasm. Among the nuclear pore complex proteins, Nup107 co-localized with Lam during this meiotic division. Surprisingly, Lam depletion resulted in a higher frequency of cytokinesis failure in male meiosis. The similar meiotic phenotype was observed in Nup107-depleted cells. Abnormal localization of Lam was found in the Nup-depleted cells at premeiotic and meiotic stages. The central spindle microtubules became abnormal and recruitment of a contractile ring component to the cleavage sites was disrupted in Lam-depleted cells and Nup107-depleted cells. Therefore, it is speculated that both proteins are required for a reinforcement of the spindle envelope, which supports the formation of central spindle microtubules essential for cytokinesis in Drosophila male meiosis.
Zhang, X., Xu, K., Wei, D., Wu, W., Yang, K. and Yuan, M. (2017). Baculovirus infection induces disruption of the nuclear lamina. Sci Rep 7(1): 7823. PubMed ID: 28798307
Summary:
Baculovirus nucleocapsids egress from the nucleus primarily via budding at the nuclear membrane. The nuclear lamina underlying the nuclear membrane represents a substantial barrier to nuclear egress. Whether the nuclear lamina undergoes disruption during baculovirus infection remains unknown. This study generated clonal cell line, Sf9-L, that stably expresses GFP-tagged Drosophila lamin B. GFP autofluorescence colocalized with immunofluorescent anti-lamin B at the nuclear rim of Sf9-L cells, indicating GFP-lamin B was incorporated into the nuclear lamina. Meanwhile, virus was able to replicate normally in Sf9-L cells. Next, alterations to the nuclear lamina were investigated during baculovirus infection in Sf9-L cells. A portion of GFP-lamin B localized diffusely at the nuclear rim, and some GFP-lamin B was redistributed within the nucleus during the late phase of infection, suggesting the nuclear lamina was partially disrupted. Immunoelectron microscopy revealed associations between GFP-lamin B and the edges of the electron-dense stromal mattes of the virogenic stroma, intranuclear microvesicles, and ODV envelopes and nucleocapsids within the nucleus, indicating the release of some GFP-lamin B from the nuclear lamina. Additionally, GFP-lamin B phosphorylation increased upon infection. Based on these data, baculovirus infection induced lamin B phosphorylation and disruption of the nuclear lamina.

BIOLOGICAL OVERVIEW

Nuclear lamins belong to the intermediate filament (IF) superfamily of proteins that includes type I and II IFs called keratins (a component of wool fibers), type III IF (such a vimentin, desmin and peripherin), type IV IF (expressed in axons, dendrites and perikarya), and type V IF, proteins making up nuclear lamina (Fuchs, 1994). Drosophila Lamin is one of two nuclear lamins. Lamin is expressed constitutively, in contrast to the second Drosophila lamin, Lamin C, which is developmentally regulated (Riemer, 1995). Lamins are the major structural proteins of the nuclear lamina, a structure that lines the nucleoplasmic surface of the inner nuclear membrane in higher eukaryotic cells. The nuclear lamina is composed of a meshwork of 10 nm filaments that are thought to provide a skeletal support for the nuclear envelope and to mediate the attachment of the nuclear envelope to interphase chromatin. Additional functions of the nuclear lamina may include the proper organization and anchoring of nuclear pore complexes. During mitosis the lamins also play a crucial role in the disassembly and reassembly of the nuclear envelope (Lenz-Bohme, 1997 and references).

Before describing the biological properties of Drosophila Lamin, the general properties of the IF superfamily members will be described. IF proteins are all predicted to share a common secondary structure. All IF proteins have a central alpha-helical domain, the rod, which is flanked by nonhelical head (amino-end) and tail (carboxy-end) domains. The rods of two polypeptide chains intertwine in a coiled-coil fashion. Throughout the alpha-helical sequences are repeats of hydrophobic amino acids, such that the first and fourth repeats of every seven residues are frequently apolar. This provides a hydrophobic seal on the helical surface, enabling the coiling between two IF polypeptides. The IF alpha-helical rod is subdivided by three short nonhelical linker segments, which often contain proline or multiple glycine residues (Fuchs, 1994).

For all IFs, the first step in assembly is the formation of parallel, in-register dimers. Upon assembly, lamin dimers align in a head-to-tail fashion to form linear polymers. Most IF proteins can form functional homodimers. Each 10-nm IF filament is composed of smaller protofibrils. IFs appear to have approximately four protofibrils per unit width. It is thought that the conserved amino ends of IF proteins play an important role in assembly of the 10-Nm filament structure. One role of IF tails may be to control lateral associations at the protofilament and protofibril level, thereby influencing filament diameter. In addition, the IF head might promote lateral associations of protofilaments and protofibrils. It is known that headless lamins cannot form a linear array of dimers typical of tail-less and wild-type lamins, suggesting that the lamin head might function in tetramer elongation. Phosphorylation is known to negatively regulate IF assembly. It is thought that the archetypal IF gene had a laminlike structure (Fuchs, 1994 and references).

Lamin Dm0, the precursor form of Lamin, has an apparent molecular mass of 76 kDa and is rapidly processed proteolytically in the cytoplasm into a form migrating at 74 kDa (Lamin Dm1). Lamin Dm1 is imported into the nucleus, where about 50% is posttranslationally modified into a slower migrating form (75 kD) called Lamin Dm2. In vivo pulse-chase studies indicated that lamins Dm1 and Dm2 are in equilibrium. Treatment of lamins Dm1 and Dm2 with phosphatase results in a single form that comigrates with Lamin Dm1 (Smith, 1987 and Smith, 1989). Lamins Dm1 and Dm2 are present as a random mixture of homo- and heterodimers. It is thought that serine 25 is the Lamin Dm2-specific phosphorylation site (Stuurman, 1995).

Lamin is known to interact directly with highly conserved sequences of DNA. Lamin binds with high affinity to scaffold/matrix-associated regions (M/SARs). These DNA sequences are held responsible for mediating the interaction between the nuclear matrix and chromatin. M/SARs are several hundred base pairs long and contain stretches of AT-rich sequences that are likely to form an open chromatin configuration. Indeed, the binding of M/SARs to lamin polymers involves single-stranded regions. In addition, this binding is saturable and requires the minor groove. Lamin polymers also bind to Drosophila centromeric and telomeric sequences. The polymerized alpha-helical rod domain of Lamin, on its own, provides for specific binding to the fushi tarazu M/SAR (Zhao, 1996). The ftz M/SAR functions as an autonomously replicating sequence (ARS) in the budding yeast S. cerevisiae. This M/SAR is found in a 2.57 kb ftz upstream regulatory element. A 189 base pair minimal fragment has ARS function. However, based on growth rates and mitotic stability, its activity is lower than that of the entire SAR. The addition of flanking sequences, including as little as 100 bp of AT-rich DNA to the left of the minimal sequence, can enhance the replicative ability of the ARS. These results implicate lamins in initiation of DNA replication (Amati, 1990 a and b)

Several proteins are associated with the nuclear lamina, and specifically with Lamin proteins. The interaction of lamins with the inner nuclear membrane may be supported by integral membrane proteins, e.g., the putative lamin receptor p54 (Bailer, 1991) or LAPs, the lamina-associate proteins (Foisner, 1993).

One protein associated with Drosophila Lamin is Otefin. Otefin is the corruption and transliteration of the Hebrew word "otef," meaning envelope. Otefin is a peripheral protein of the inner nuclear membrane in Drosophila. During nuclear assembly in vitro, it is required for the attachment of membrane vesicles to chromatin. Otefin colocalizes with Lamin derivatives in situ and presumably in vivo and is present in all somatic cells examined during the different stages of Drosophila development. Otefin is a phosphoprotein in vivo and is a substrate for in vitro phosphorylation by cdc2 kinase and cyclic AMP-dependent protein kinase. It is suggested that Otefin plays a role in the assembly of the Drosophila nuclear envelope (Ashery-Padan, 1997b).

The fs(1)Ya protein (Ya stands for "young arrest") is an essential, maternally encoded, nuclear lamina protein that is under both developmental and cell cycle control. A strong Ya mutation results in early arrest of embryos. Ya mutant embryos arrest with abnormal nuclear envelopes prior to the first mitotic division. Ya unfertilized eggs contain nuclei of different sizes and condensation states, apparently due to abnormal fusion of the meiotic products immediately after meiosis. Lamin is localized at the periphery of the uncondensed nuclei in these eggs. These results suggest that Ya function is required during and after egg maturation to facilitate proper chromatin condensation, rather than to allow a lamin-containing nuclear envelope to form. Ya might bind to chromatin and organize the chromatin structure in early embryos in a way that permits DNA replication. Whether Ya functions in conjunction with lamin is unknown (Liu, 1995).

A Drosophila Lamin mutant shows a severe phenotype: this includes retardation in development, reduced viability, sterility, and impaired locomotion. Mutant adult flies die within 2 weeks after eclosion. Late stages of oogenesis are rarely detected in mutant ovaries, and the egg chambers present show an abnormal morphology. In heads from homozygous mutant flies, Lamin is significantly decreased in nuclei of the densely packed cell bodies of the central nervous system. Reduced Lamin expression causes an enrichment of nuclear pore complexes in cytoplasmic annulate lamellae and in nuclear envelope clusters. Annulate lamellae are stacked sheets of membranes in the cytoplasm that contain pore complexes in high densities and are often continuous with rough endoplasmic reticulum in several cells, particularly the densely packed somata of the central nervous system. Defective nuclear envelopes are also observed. The lack of full lethality at early developmental stages may be due to maternal transmission from heterozygous mothers. Indeed, Lamin protein is highly enriched inside the oocyte nucleus, which may serve as a storage compartment for lamin required during the early nuclear divisions in the embryo. These data constitute the first genetic proof that lamins are essential for the structural organization of the cell nucleus (Lenz-Bohme, 1997).

The B-type lamin is required for somatic repression of testis-specific gene clusters

Large clusters of coexpressed tissue-specific genes are abundant on chromosomes of diverse species. The genes coordinately misexpressed in diverse diseases are also found in similar clusters, suggesting that evolutionarily conserved mechanisms regulate expression of large multigenic regions both in normal development and in its pathological disruptions. Studies on individual loci suggest that silent clusters of coregulated genes are embedded in repressed chromatin domains, often localized to the nuclear periphery. To test this model at the genome-wide scale, transcriptional regulation of large testis-specific gene clusters was studied in somatic tissues of Drosophila. These gene clusters showed a drastic paucity of known expressed transgene insertions, indicating that they indeed are embedded in repressed chromatin. Bioinformatics analysis suggested the major role for the B-type lamin, LamDm(o), in repression of large testis-specific gene clusters, showing that in somatic cells as many as three-quarters of these clusters interact with LamDm(o). Ablation of LamDm(o) by using mutants and RNAi led to detachment of testis-specific clusters from nuclear envelope and to their selective transcriptional up-regulation in somatic cells, thus providing the first direct evidence for involvement of the B-type lamin in tissue-specific gene repression. Finally, it was found that transcriptional activation of the lamina-bound testis-specific gene cluster in male germ line is coupled with its translocation away from the nuclear envelope. These studies, which directly link nuclear architecture with coordinated regulation of tissue-specific genes, advance understanding of the mechanisms underlying both normal cell differentiation and developmental disorders caused by lesions in the B-type lamins and interacting proteins (Shevelyov, 2009).

It was hypothesized that, in addition to the somatic silencing of testis-specific gene clusters, LamDm0 is also required for attachment of these clusters to the nuclear envelope. To test this hypothesis, intranuclear positions of the 60D1 and 22A1 regions was determined in the interphase nuclei of cultured S2 cells in which LamDm0 was ablated by RNAi. Fluorescence in situ hybridization (FISH) combined with immunostaining for LamDm0 confirmed RNAi-induced ablation of LamDm0, and showed approximately 2-fold decrease in frequency of the 60D1 and 22A1 loci bound to lamina. Next, association of the 60D1 gene cluster with nuclear envelope wa analyzed during male germ-line differentiation. The whole-mount testes dissected from the third instar larvae were analyzed for intranuclear localization of the 60D1 region by FISH combined with immunostaining for LamDm0. Morphologically, a group of small cells is located at the end of testis and includes spermatogonia, cyst cells and stem cells, in which the 60D1 locus is silent. In these cells, the 60D1 region is associated with nuclear lamina in 76% of nuclei, similarly to the cultured somatic S2 cells. On the contrary, in spermatocytes (identified by characteristic large nuclei), in which testis-specific genes in the 60D1 region are expressed, this region is found at the nuclear envelope in only 6% of the nuclei. Thus, transcriptional activation of the testis-specific gene cluster in male germ line is coupled to its dissociation from the nuclear envelope. Similarly, detachment from the nuclear lamina has been associated with transcriptional activation of other lamina-bound loci both in Drosophila and in mammals. These observations strongly suggest a model in which gene repression is controlled in a cell type-specific manner through regulated tethering of chromatin to the nuclear lamina. Further dissection of the mechanisms that mediate repression of lamina-bound multigenic regions and control localization of these regions at nuclear envelope will provide new insights into coordinated regulation of tissue-specific genes, thus advancing understanding of cell differentiation both in normal development and in disease (Shevelyov, 2009).

Role of histone deacetylases in gene regulation at nuclear lamina

Theoretical models suggest that gene silencing at the nuclear periphery may involve 'closing' of chromatin by transcriptional repressors, such as histone deacetylases (HDACs). This study provides experimental evidence confirming these predictions. Histone acetylation, chromatin compactness, and gene repression in lamina-interacting multigenic chromatin domains were analyzed in Drosophila S2 cells in which B-type lamin, diverse HDACs, and lamina-associated proteins were downregulated by dsRNA. Lamin depletion resulted in decreased compactness of the repressed multigenic domain associated with its detachment from the lamina and enhanced histone acetylation. The data reveal the major role for HDAC1 in mediating deacetylation, chromatin compaction, and gene silencing in the multigenic domain, and an auxiliary role for HDAC3 that is required for retention of the domain at the lamina. These findings demonstrate the manifold and central involvement of class I HDACs in regulation of lamina-associated genes, illuminating a mechanism by which these enzymes can orchestrate normal and pathological development (Milon, 2012).

This study provides direct experimental evidence for the long-persisting assumptions that HDACs are involved in gene silencing at the nuclear lamina, by identifying Class I enzymes HDAC1 and HDAC3 as the major players in this mechanism. Likewise gene silencing, histone hypoacetylation and chromatin compaction in the multigenic chromatin domain are lamin-dependent. Moreover, HDAC1 was identified as the key factor required for silencing and specifically responsible for histone H4 deacetylation, and the data implicate HDAC3 as an auxiliary factor specifically responsible for localization of the repressed chromatin at the lamina. The 'closed' chromatin configuration of the repressed domain also depends on HDAC1 and thus probably mediates the major repressive action of this enzyme at the nuclear periphery. Published data indicate that the 60D1 cluster interacts with HDAC1, in particular in the Crtp and Pros28.1B regions, supporting direct involvement of this enzyme in histone deacetylation. A model is proposed in which Class I HDACs participate in lamina-dependent gene silencing through diverse pathways: HDAC1, tethered to the lamin scaffold by LEM domain proteins, is involved in deacetylation of histones H3 and H4 and 'closing' of lamina-bound chromatin while HDAC3 contributes to histone H3 deacetylation and retention of the repressed chromatin at the lamina. Interestingly, a recent study showed that HDAC3 is also involved in peripheral localization of the lamina-interacting chromatin in mammals (Zullo, 2012) indicating that this mechanism is conserved between diverse animals (Milon, 2012).

Lamina-associated chromatin domains harbor numerous cell type-specific genes that must be precisely regulated to orchestrate cell differentiation and development. Genetic defects in the lamina components result in severe and currently incurable tissue degenerative disorders known as laminopathies. Identification of the key role of Class I HDACs, and particularly HDAC1, in lamina-associated gene silencing implies that modulation of this enzyme may help to restore gene expression disrupted by nuclear lamina defects, and may be instrumental in establishing new expression patterns in pluripotent cells to guide their differentiation (Milon, 2012).


GENE STRUCTURE

The Drosophila Lamin gene is developmentally regulated, giving rise to a 2.8 kb maternal transcript and a 3.0 kb zygotic transcript. The different transcripts are generated by utilizing different polyadenylation sites. None of the putative lamin polyadenylation signals contains the consensus AAUAA sequence. The choice between the different polyadenylation signals might depend on maternal fators that more efficiently recognize the polyadenylation signal of the 2.8 kb transcript (Osman, 1990 and Gruenbaum, 1988)

Bases in 5' UTR - 147

Exons - 4

Bases in 3' UTR - 710 (maternal) and 921 (zygotic)


PROTEIN STRUCTURE

Amino Acids - 621

Structural Domains

Highly specific features of lamins include a nuclear localization signal, a C-terminal CaaX sequence (where C=cysteine; a=aliphatic amino acid; X=any amino acid) and characteristic phosphorylation sites in the N-terminal head and C-terminal tail domains. The nuclear localization signal is responsible for rapid transport of lamins into the nucleus, thus preventing cytoplasmic assembly. Modification by isoprenylation and carboxymethylation at the CaaX motif targets lamins to the inner nuclear membrane (Lenz-Bohme, 1997 and references).


Lamin: Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References

date revised: 16 July 97  

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