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PMID: 14727112 Published · ppublish English Comparative Study Journal Article Research Support, U.S. Gov't, Non-P.H.S.

Biochemical and immunological characterization of pea nuclear intermediate filament proteins.

Planta ·Vol. 218 ·No. 6 ·2004-04-00 ·Pages 965-75

Blumenthal SS, Clark GB, Roux SJ

Abstract

In immunoblot assays, at least three putative nuclear intermediate filament (NIF) proteins were detected in nuclear envelope-matrix (NEM) and lamin (L1) fractions of nuclei from plumules of dark-grown pea (Pisum sativum L.) seedlings. These NIF proteins had apparent molecular masses of ca. 65, 60, and 54 kDa (also referred to as p65, p60, and p54), and appeared as multiple isoelectric forms, with pIs ranging from ca. 4.8 to 6.0. Polyclonal and monoclonal antibodies were raised to the 65-kDa NIF protein bands excised from gels after electrophoresis. These anti-pea antibodies were specifically cross-reactive with the pea nuclear p65, p60, and p54 proteins and also with chicken lamins. Sequence alignment of peptide fragments obtained from the 65- and 60-kDa pea NIF proteins showed similarity with animal intermediate filament proteins such as lamins and keratins and with certain plant proteins predicted to have long coiled-coil domains. These pea NIF proteins were further purified and enriched from the NEM fraction using methods similar to those used for isolating animal lamins. When negatively stained and viewed by transmission electron microscopy, the filaments in the pea lamin (L1) fraction appeared to be 6-12 nm in diameter. As assayed by immunofluorescence cytochemistry using a confocal laser-scanning microscope, fixed pea plumule cells displayed uniform as opposed to peripheral nuclear staining by several of the antibody preparations, both polyclonal and monoclonal. This report describes the biochemical and immunological properties of these pea NIF proteins.

Keywords
NASA Discipline Plant Biology Non-NASA Center
MeSH Terms
Amino Acid Sequence Antibodies, Monoclonal/immunology Antibody Specificity Intermediate Filament Proteins/chemistry,immunology,ultrastructure Microscopy, Electron Molecular Sequence Data Nuclear Envelope/immunology,metabolism Peas/metabolism Plant Proteins/chemistry,immunology Sequence Alignment Sequence Homology, Amino Acid
Chemicals
Antibodies, Monoclonal Intermediate Filament Proteins Plant Proteins
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Blumenthal Sonal S D
School of Biological Sciences, Section of Molecular Cell and Developmental Biology, The University of Texas, Austin, TX 78712, USA.
Clark Gregory B
Roux Stanley J
Investigators
1 investigators, click to expand
Roux S J
U TX, Austin
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40 references, click to expand
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Article Info
Journal
Planta
Abbr.
Planta
ISSN
0032-0935
DOI
10.1007/s00425-003-1182-5
Published
2004-04-00
Epub
2004-00-15
Pages
965-75
Language
English
Region
Germany
NLM ID
1250576
Subset
IM
External Links
PubMed source DOI: 10.1007/s00425-003-1182-5 View journal details
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