Karyophilic peptides: Applications to the study of nuclear transport

Goldfarb, David S.

doi:10.1016/0309-1651(88)90090-2

Cited by 30 publications

(9 citation statements)

References 38 publications

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“…Both proteins, like p140, are not detectable in the cytoplasm by indirect immunofluorescence microsco W or on Western blots after subcellular fraetionation. In addition, No38 was found to bind to immobilized wild-type NLS peptides (Goldfarb, 1988). In this case, however, the absence of binding to the mutant NLS peptide was not tested.…”

Section: Discussionmentioning

confidence: 96%

A nuclear localization signal binding protein in the nucleolus.

Meier

Blobel

1990

The Journal of Cell Biology

154

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Abstract. We used functional wild-type and mutant synthetic nuclear localization signal peptides of SV-40 T antigen cross-linked to human serum albumin (pepfide conjugates) to assay their binding to proteins of rat liver nuclei on Western blots. Proteins of 140 and 55 kD (p140 and p55) were exclusively recognized by wild-type peptide conjugates. Free wild-type peptides competed for the wild-type peptide conjugate binding to p140 and p55 whereas free mutant peptides, which differed by a single amino acid from the wild type, competed less efficiently. The two proteins were extractable from nuclei by either low or high ionic strength buffers. We purified p140 and raised polyclonal antibodies in chicken against the protein excised from polyacrylamide gels. The anti-pl40 antibodies were monospecific as judged by their reactivity with a single nuclear protein band of 140 kD on Western blots of subcellular fractions of whole cells. Indirect immunofluorescence microscopy on fixed and permeabilized Buffalo rat liver (BRL) cells with anti-pl40 antibodies exhibited a distinct punctate nucleolar staining. Rhodamine-labeled wild-type peptide conjugates also bound to nucleoli in a similar pattern on fixed and permeabilized BRL cells. Based on biochemical characterization, p140 is a novel nucleolar protein. It is possible that p140 shuttles between the nucleolus and the cytoplasm and functions as a nuclear import carrier.

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Section: Discussionmentioning

confidence: 96%

A nuclear localization signal binding protein in the nucleolus.

Meier

Blobel

1990

The Journal of Cell Biology

154

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“…Both the association of karyophilic substrates with the nuclear envelope and their rate of accumulation in nuclei are influenced by the multiplicity of signal sequences displayed by individual substrate molecules (ll, 15, 31). We have observed both specific radial binding to the NPC transporter and nuclear import of 60-A gold particles stabilized with BSA coupled to high numbers of mutant peptides, which contain a threonine as replacement for lysine-128 in the SV-40 large T antigen signal sequence; a low multiplicity of peptides resulted in poor transport (15,23). Therefore, multivalent effects may play a role in the binding and translocation phases of nuclear import.…”

Section: Peripheral Binding To the Npc Transportermentioning

confidence: 97%

“…The translocation step is shown schematically as a dilation of the transporter; however, the actual mechanism is not known. transport (15,23). Therefore, multivalent effects may play a role in the binding and translocation phases of nuclear import.…”

Section: Peripheral Binding To the Npc Transportermentioning

confidence: 99%

Protein import through the nuclear pore complex is a multistep process.

Akey

Goldfarb

1989

The Journal of Cell Biology

209

127

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The transport of macromolecules across the nuclear envelope is mediated by the nuclear pore complex (NPC). Using cryo-electron microscopy and image processing we have mapped the interaction of three specific gold probes with the NPC and obtained projection maps of two possible intermediates in nuclear import. The probes used in these experiments were (a) mAb-414, which cross-reacts with Xenopus nucleoporins containing O-linked N-acetyl glucosamines; (b) wheat germ agglutinin, a transport inhibitor; and (c) nucleoplasmin, a transport substrate. Strong binding sites of the three probes are circularly arrayed on NPCs between radii of 100 and 125 A and may be coextensive. These results suggest that nucleoplasmin-gold (NP-gold) can form at least three distinct complexes with a central transport assembly of the NPC, which may represent intermediates of a multistep protein import pathway. Initially, NP-gold appears to bind at multiple sites located around the periphery of the closed NPC transporter and also directly over the center where it can dock. In a subsequent step NP-gold is translocated through the nuclear pore.

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“…Several NBPs have been identified, including the 60 kDa, 70 kDa, and Noppl40 proteins from rat liver (Adam et al, 1989;Meier & Blobel, 1992), the B23/No38 from mouse (Goldfarb, 1988), and the NSRl protein from yeast (Lee, Xue & Melese, 1991). The B23/No38 and NoppUO shuttle between the nucleus and cytoplasm and such shuttling, in general, is suggested to play important roles in nucleocytoplasmic transport (Borer et al, 1989;Meier & Blobel, 1992).…”

Section: A Surface Viewmentioning

confidence: 99%

Plasmodesmata and the supracellular nature of plants

1993

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SUMMARY In the classical formulation of Münch (1930), plasmodesmata are considered to form simple cytoplasmic bridges between neighbouring plant cells to create the symplasm. This concept has dominated, if not monopolized, the thinking of plant biologists and in particular plant physiologists over the last few decades. Recent advances in ultrastructural, physiological and molecular studies on plasmodesmata indicate that this simple view is in need of revision. Structurally, the higher plant plasmodesma has been revealed to be a supramolecular complex consisting of membranes and proteins. Functionally, evidence is at hand that this complex structure appears to have evolved not only to control the size exclusion limit for intercellular diffusion of metabolites and small molecules, but also to potentiate and regulate intercellular trafficking of macromolecules, including proteins and nucleic acids. In this regard, plasmodesmal transport may share parallel regulatory mechanisms with nucleocytoplasmic transport. Based on these findings, we advance the hypothesis that plants function as supracellular, rather than multicellular, organisms. As such, the dynamics of the plant body, including cell differentiation, tissue formation, organogenesis and specialized physiological function(s), is subject to plasmodesmal regulation. Plasmodesmata presumably accomplish such regulatory roles by trafficking informational molecules which orchestrate both metabolic activity and gene expression. Current and future studies on the evolutionary origin(s) of plasmodesmata are likely to provide valuable information in terms of the genetic and molecular basis for the supracellular nature of plants.

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Karyophilic peptides: Applications to the study of nuclear transport

Cited by 30 publications

References 38 publications

A nuclear localization signal binding protein in the nucleolus.

A nuclear localization signal binding protein in the nucleolus.

Protein import through the nuclear pore complex is a multistep process.

Plasmodesmata and the supracellular nature of plants

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