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From CellBiology

Lamin B1

Cellular localization of LAP2alpha, DNA and Lamin B in NRK Cells (A) and Frozen Rat Liver cells (B).[1]

Lamin B1 is a specific 65–68 kDa fibrous protein that is a subset of a larger group of lamin proteins. Lamins are polypeptides classified as V-type Intermediate Filaments with many nuclear functions. The localization of lamins is within the underlying nuclear lamina of the nuclear envelope and provides structural support and integrity. Human cells contain two dominant lamins types classified into either A-type or B-type dependent on biochemical properties, expression patterns, sequence homology and mitotic behaviours.[2][3][4][5]

Lamin B1 is graded into the Type-B lamin category and is activated from cellular development onwards for nuclear envelope construction. On the contrary, Type-A lamins are predominantly found as insoluble proteins during cellular development and activated during cellular differentiation. Lamin B1 comprises the majority of lamin species within the nuclear lamina and provides an intermediate linkage by anchoring at sites of chromatin domains and the nuclear envelope. It is also apparent during mitosis that Type-B lamins differs from Type-A through the absent display of solubility properties. Additionally, there is an alternate exhibition of methods and sequencing of chromatin association during mitosis.[6][7]


Structure

Conserved lamin structure.[2]
Generalized structure of cytoplasmic intermediate-filament proteins compared to lamins.[8]

The structure of lamins is conserved across all species including lamin B1. It involves a central coiled rod region associated with non-helical nitrogen terminal and carbon terminal domains at either end of the rod.[4][9]


  • The central coiled α-helical rod domain contains approximately 356 residues in a quasiheptad repeat sequence motif formation with a conserved secondary structure. It is discovered as two strands of α-helical coiled coil structures in a parallel spiral alignment around a common axis. This rod region is called an inregistered dimer and is involved in regulation of lamin dimerisation with the head and tail domains controlling polymerization and higher-order assembly.[4][9]


  • The carbon terminal domain may be segregated into three components that are a nuclear localization signal (NLS), an immunoglobulin (Ig) and a CAAX motif. The ‘CAAX’ motif involves the ‘A’ regions being aliphatic side chains, ‘C’ being a cysteine residue and ‘X’ being any residue.[9]


Formation

Development and formation of lamin B1 involves transcription and translation of the specific gene, LMNB1. Supplementary maturation stages of lamin B1 are needed to render the protein functional that include post-translational processing at the region of the CAAX motif on the C terminus.[2][5][7][10]


The maturation process begins with the cysteine component of the CAAX motif required to undertake farnesylation, followed by, cleavage of AAX by an endopeptidase. Subsequently, the remaining cysteine residue undergoes carboxymethylation creating the mature form of lamin B1. The developmental process of farnesylation and endoproteolysis aids immature lamin B1 by providing stability within the architecture of the nuclear envelope.[10][11]


Lamins undertake moderately analogous developmental pathways although differ in the specific gene expressed in its development. As lamin B1 is derived from LMNB1 gene expression, lamin B2 develops almost identically but rather is encoded by the LMNB2 gene. In addition, Type-A lamins are encoded by a singular LMNA gene creating four possible lamin isoforms. The two primary products are lamin A and C whilst minor products include lamin A∆10 and germ cell-specific lamin C2.[2][10][11]


Function

Co-localization of LAP2alpha, lamin B and lamin A/C at early stages of Nuclear Envelope assembly.[1]

Lamin B1 is required during embryonic organogenesis onwards through the provision of nuclear envelope integrity and chromosomal segregation in mitosis. Furthermore, this protein is integral in embryonic development of bones and lungs along with construction of nuclei and cellular growth and differention. Lamin B1 also provides adherence of the nuclear lamina onto the nuclear envelope and is required for positioning of chromosome territories and distribution of chromatin.[3][6][10]


Vergnes et al. (2004) undertook an experimental study in HeLa cells of mice using mutant lamin B1 species and to observe the primary functions of lamin B1 within the nucleus. In the experiment, modifications were introduced to lamin B1 species to induce mutations as abnormal lamin B1 species are not exhibited in normal cells. Expression of mutant lamin B1 caused detrimental impacts on normal lung development with assorted cases of respiratory failures and perinatal lethality. In addition, modifications of the nuclear components were evident within affected cells through an irregular shaped nuclear envelope and abnormal chromatin distribution. In this case, evidence suggests that a reduction of functional lamin B1 species triggers a compensative response of lamin B2 species in attempt to sustain normal nuclear envelope functions during embryonic development.[6]


Moreover, specific structural changes induced by physical disruption within lamin B1 reduced its capability in fulfilling its normal tasks. The removal of the carboxyl terminal end of lamin B1 caused disruptions to chromatin-binding regions within the lamin species whilst the absence of the specific carboxyl-terminal CAAX motif led to inefficient targeting of Type-A lamins.[6]


Abnormalities

Adult-onset autosomal dominant leukodystrophy (ADLD) is characterized by genomic duplication of lamin B1 within cells and remains the only human lamin B1-associated disease. ADLD is a demyelinating neurological disorder affecting the central nervous system through gradual reduction of myelin levels over time. It involves cells carrying an additional gene that encodes lamin B1 and when over-expressed, increases gene dosage within brain tissue. As a result, progressive alterations occur that are supported by the toxic environment generated by accumulated lamin B1 proteins and greater cellular malleability in ageing cells.[12]


Current research

Recent research undertaken by Tang, et al (2008) involved conducting experiments using HeLa cells to demonstrate the impacts of reduced expression of lamin B1. It has been concluded that limited lamin B1 activity indicated a loss of cellular RNA synthesis with prolonged reduction of expression leading to adverse effects promoting apoptosis.[7][13]


The apparent stoppage of RNA synthesis was halted by inactivity of polymerase II and polymerase I in the transcription process attributed by low lamin B1 levels. The effects of reduced lamin B1 expression led to expansion of inter-chromatin domains and associated structural and spatial modifications of chromosomal territories towards the nuclear periphery. In addition, nuclear chromosomal territory repositioning and collapse of related chromatin was present amongst the nuclear structural changes. Therefore, lamin B1 contribution within the nuclear cytoskeleton remains vital towards maintaining regular RNA synthesis and the global nuclear architecture of cells.[11][13]


Click here for a general overview of the nucleus


References

  1. 1.0 1.1 Dechat, T., Gotzmann, J., Stockinger, A., et al. (1998). Detergent-salt resistance of LAP2alpha in interphase nuclei and phosphorylation-dependent association with chromosomes early in nuclear assembly implies functions in nuclear structure dynamics. The EMBO journal 17(16):4887-902. PMID: 9707448
  2. 2.0 2.1 2.2 2.3 Hill, Mark (2009). 2009 Lecture 10 - Cellbiology. Retrieved May 05, 2009, from Cell Biology Wiki Web site: http://cellbiology.med.unsw.edu.au/cellbiology/index.php?title=2009_Lecture_10
  3. 3.0 3.1 Aebi, U., Cohn, J., Buhle, L., et al. (1986) The nuclear lamina is a meshwork of intermediate-type filaments. Nature 323(6088):560-4. PMID: 3762708
  4. 4.0 4.1 4.2 Hill, Mark (2009). 2009 Lecture 4 - Cellbiology. Retrieved May 10, 2009, from Cell Biology Wiki Web site: http://cellbiology.med.unsw.edu.au/cellbiology/index.php?title=2009_Lecture_4
  5. 5.0 5.1 Goldman, R.D., Gruenbaum, Y., Moir, R.D., et al. (2002) Nuclear lamins: building blocks of nuclear architecture. Genes and Development 16 (5):533-47. PMID: 11877373
  6. 6.0 6.1 6.2 6.3 Vergnes, L., Péterfy, M., Bergo, M.O., et al. (2004) Lamin B1 is required for mouse development and nuclear integrity. Proceedings of the National Academy of Sciences of the United States of America 101 (28):10428-33. PMID: 15232008
  7. 7.0 7.1 7.2 Margalit, A., Vlcek, S., Gruenbaum, Y., et al. (2005) Breaking and making of the nuclear envelope. Journal of Cellular Biochemistry 95 (3): 454-65. PMID: 15832341
  8. Hutchison, C.J., Worman, H.J. (2004). A-type lamins: guardians of the soma? Nature Cell Biology 6(11):1062-7 PMID: 15517000
  9. 9.0 9.1 9.2 Stewart, M. (1993) Intermediate filament structure and assembly. Current Opinion in Cell Biology 5(1):3-11. PMID: 8448027
  10. 10.0 10.1 10.2 10.3 Dahl, K.N., Ribeiro, A.J., Lammerding, J. (2008) Nuclear shape, mechanics, and mechanotransduction. Circulation Research 102 (11): 1307-18. PMID: 18535268
  11. 11.0 11.1 11.2 Malhas, A., Lee, C.F., Sanders, R., et al. (2007) Defects in lamin B1 expression or processing affect interphase chromosome position and gene expression. The Journal of Cell Biology 176 (5):593-603. PMID: 17312019
  12. Padiath, Q.S., Saigoh, K., Schiffmann, R., et al. (2006) Lamin B1 duplications cause autosomal dominant leukodystrophy. Nature Genetics 38 (10):1114-23 PMID: 16951681
  13. 13.0 13.1 Tang, C.W., Maya-Mendoza, A., Martin, C., et al. (2008) The integrity of a lamin-B1-dependent nucleoskeleton is a fundamental determinant of RNA synthesis in human cells. Journal of Cell Science 121 (Pt 7):1014-24. PMID: 18334554


ANAT3231 Cell Biology Homework

Lecture 4 - Nucleus

I learnt that the nuclear envelope contains various structures called nuclear pore complexes that permit the free flow of small molecules by passive diffusion and controls flow of macromolecules by facilitated diffusion.

Lecture 5 - Exocytotsis

I discovered that exocytosis to be quite straight forward although I am having difficulty understanding whether the Golgi Apparatus curves away or around the nucleus? I am also not sure whether there is an answer to this question.

Lecture 7 - Mitochondria

Mitochondria is important for cell signalling, cell differentiation and apoptosis. Cellular movement requires significant energy from mitochondria through muscle contractions along with cell motility through cilia, flagella and amoeboid actions. photosynthesis also requires a great deal of mitochondria.

Lecture 8 - Adhesion

CAM stands for cell adhesion molecule

  • Ng-CAM is Neuroglia Cell Adhesion Molecule
  • I-CAM is Intercellular Cellular Adhesion Molecule
  • L-CAM is Liver Cell Adhesion Molecule

Lecture 10 - Intermediate Filaments

The layer of tissue located between the basal and granulosa layers is called stratum spinosum. Intermediate Filaments are located within desmosomes on the basolateral membranes of the cells for intercellular tranport of molecules.

Laboratory 6 - Cytoskeleton Exercise

  • Genotype A: Tm4 over-expressing B35 cells
  • Genotype B: wt (wild type: lack of expression) control B35 cells


There are differences in the changes through the phenotypes created due to the expression of Tm4.

  • The Tm4 over-expressing B35 cells experienced a greater level of production of neurites and lamella with increasing number of processes with a maximum count of 6 processes in some cells.
  • The control wild-type B35 cells appear to have a greater extension of thier processes with a reduction in diameter. Theses processes are modified into a opposing manner that protrude 180 degrees to each other along the same axis.

I hypothesise that the over-expression of Tm4 triggers the growth of neurites and lamella in the actin cell cortex at the expense of extending pre-existing processes within cells.

Lecture 14 - Extracellular Matrix 2

There are two forms of generating confocal microscopy through the use of either a:

  • Laser Scanning Confocal Microscope; or
  • Spinning Disc Confocal Microscope

Lecture 15 - Cell Cycle

The S phase during the cell cycle is the abbreviated version of 'sythesis phase' that occurs during interphase between G1 Phase and G2 Phase