DOI Seite / Zitierlink:
https://doi.org/10.11588/diglit.48120#0083
Abstracts
65
Isolation and Sequencing of Human Lamin ii cDNA: Mammalian
Lamins A, B and C Contain Leucine Heptad Repeats
K. M. Pollard, E.K1. Chan, B. J. Grant, K.F. Sullivan, E. M. Tan, and C.A. Glass
WM Keck Autoimmune Disease Center and DNA Coro Facility, Department of Molecular
and Experimental Medicine and Department of Molecular Biology, Research Institute of
Scripps Clinic, La Jolla, CA 92037, USA
The nuclear lamins A, B and C form a meshwork of filaments on the inner,
or nucleoplasmic, surface of the nuclear envelope. Autoantibodies to all three
mammalian lamins have been found in the serum of patients with certain im-
munological disorders such as systemic lupus erythematosus, scleroderma and
autoimmune liver disease. These sera have proven useful reagents for the
characterisation of lamins in mammalian and other species and for the cloning
of human lamins A and C. In this report we describe our use of a NZB/W Fl
mouse monoclonal autoantibody, called 72B9, to clone nuclear lamin B using a
Agtll cDNA expression library prepared from the human leukemia T cell line
MOLTA. A unique feature of this study is that the monoclonal antibody used
reacts in immunofluorescence studies with the nucleolus and targets the 34 kDa
protein component of the nucleolus restricted U 3 small nuclear ribonucleopro-
tein particle (U3snRNP). The identity of the encoded protein as lamin B was
established by both biochemical and immunological criteria which included im-
munoprecipitation of in vitro translated product of RNA transcripts of the cDNA
by lamin B specific antibodies. Detection of mRNA in RL60 cells, that express
only lamin B, or HeLa cells, that express all three major lamins A, B and C, and
the colocalisation of in vitro translated product with HeLa cell lamin B by two-
dimensional gel electrophoresis, strongly suggests that a single lamin B is express-
ed in mammalian somatic cells. Southern blot analysis revealed that a single gene
encodes human lamin B. Inspection of the deduced amino acid sequence of lamin
B revealed the presence in coil 1 B of the a-helical domain of a leucine heptad
repeat region. Leucine heptad repeats were also found in coil 1 B of other lamins,
including the type A lamins which also contain a leucine heptad repeat in coil 2
of the a-helical domain. These leucine heptad repeats are centrally located on
coils 1B and 2, which suggests a role in the dimerisation of lamin monomers. The
lamins therefore belong to the growing list of nuclear proteins whose dimerisation
domains contain leucine heptad repeats.
65
Isolation and Sequencing of Human Lamin ii cDNA: Mammalian
Lamins A, B and C Contain Leucine Heptad Repeats
K. M. Pollard, E.K1. Chan, B. J. Grant, K.F. Sullivan, E. M. Tan, and C.A. Glass
WM Keck Autoimmune Disease Center and DNA Coro Facility, Department of Molecular
and Experimental Medicine and Department of Molecular Biology, Research Institute of
Scripps Clinic, La Jolla, CA 92037, USA
The nuclear lamins A, B and C form a meshwork of filaments on the inner,
or nucleoplasmic, surface of the nuclear envelope. Autoantibodies to all three
mammalian lamins have been found in the serum of patients with certain im-
munological disorders such as systemic lupus erythematosus, scleroderma and
autoimmune liver disease. These sera have proven useful reagents for the
characterisation of lamins in mammalian and other species and for the cloning
of human lamins A and C. In this report we describe our use of a NZB/W Fl
mouse monoclonal autoantibody, called 72B9, to clone nuclear lamin B using a
Agtll cDNA expression library prepared from the human leukemia T cell line
MOLTA. A unique feature of this study is that the monoclonal antibody used
reacts in immunofluorescence studies with the nucleolus and targets the 34 kDa
protein component of the nucleolus restricted U 3 small nuclear ribonucleopro-
tein particle (U3snRNP). The identity of the encoded protein as lamin B was
established by both biochemical and immunological criteria which included im-
munoprecipitation of in vitro translated product of RNA transcripts of the cDNA
by lamin B specific antibodies. Detection of mRNA in RL60 cells, that express
only lamin B, or HeLa cells, that express all three major lamins A, B and C, and
the colocalisation of in vitro translated product with HeLa cell lamin B by two-
dimensional gel electrophoresis, strongly suggests that a single lamin B is express-
ed in mammalian somatic cells. Southern blot analysis revealed that a single gene
encodes human lamin B. Inspection of the deduced amino acid sequence of lamin
B revealed the presence in coil 1 B of the a-helical domain of a leucine heptad
repeat region. Leucine heptad repeats were also found in coil 1 B of other lamins,
including the type A lamins which also contain a leucine heptad repeat in coil 2
of the a-helical domain. These leucine heptad repeats are centrally located on
coils 1B and 2, which suggests a role in the dimerisation of lamin monomers. The
lamins therefore belong to the growing list of nuclear proteins whose dimerisation
domains contain leucine heptad repeats.