Observations on the Structure of the Nucleocapsids of some Paramyxoviruses

Finch, J.T.; Gibbs, Adrian

doi:10.1099/0022-1317-6-1-141

Cited by 85 publications

(41 citation statements)

References 13 publications

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“…Differences in the tightness of coiling and flexibility of the nucleocapsid of different paramyxoviruses examined under similar conditions have been noted in the past (1,5,7,26)-e.g., the nucleocapsid of Sendai virus has been observed to be more tightly coiled and rigid than that of SV5, Newcastle disease virus, or measles virus. We therefore investigated the effect of salt concentration on the nucleocapsid of SV5.…”

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confidence: 73%

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Conformation of the helical nucleocapsids of paramyxoviruses and vesicular stomatitis virus: reversible coiling and uncoiling induced by changes in salt concentration.

Heggeness¹,

Scheid²,

Choppin³

1980

Proc. Natl. Acad. Sci. U.S.A.

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The conformations of the helical nucleocapsids of the paramyxoviruses Sendai virus and simian virus 5, and of a rhabdovirus, vesicular stomatitis virus, have been found to vary extensively with changes in salt concentration. In 10 mM sodium phosphate buffer at pH 7.2, the nucleocapsids are loosely coiled or almost completely extended; with increasing concentrations of NaCl they become more tightly coiled and less flexible. Under isotonic conditions (150 mM) the Sendai virus nucleocapsid is moderately tightly coiled but still curved and apparently flexible, whereas at 400 mM or higher it is very tightly coiled, with the appearance of a rigid rod. These saltdependent changes in conformation were also found with nucleocapsids composed of proteolytically cleaved protein subunits. Because of the effect of salt concentration, and the fact that it may change during the preparation of negatively stained samples for electron microscopy, it was necessary to fix the nucleocapsids before negative staining to preserve their original conformation. The striking changes in nucleocapsid conformation in response to the ionic milieu indicate the plasticity of its helical structure and suggest that changes in the microenvironment of the nucleocapsid could influence its conformation during viral RNA transcription and replication or during virus assembly by budding, processes in which changes in the coiling of the nucleocapsid or its flexibility could be important. The nucleocapsids of paramyxoviruses, such as Sendai virus, simian virus 5 (SV5), and Newcastle disease virus, are single, left-handed, helical structures which, when tightly coiled, have a diameter of 15-18 nm and a length of ;1 ,m (1-5). They are composed of single-stranded RNA and -2600 viral structural protein subunits (NP) with a molecular weight of t60,000 (6, 7). Two other proteins (L and P) are associated with the nucleocapsid and thought to be involved in the viral RNA transcriptase activity (8-12). The nucleocapsids of rhabdoviruses-e.g., vesicular stomatitis (VSV) and rabies viruses-also consist of a single-stranded RNA complexed with a single major structural protein (N) and two associated proteins (NS and L) (13-18). In the VSV virion, the nucleocapsid forms a coil t40 nm in diameter but, when separated from the viral membrane, it is often found in helices of considerably smaller diameter (13-17).The conformation of paramyxovirus nucleocapsids has shown wide variations as observed in the electron microscope. They have been seen to have tightly stacked turns and the overall appearance of relatively stiff rods ;1 ,um in length and also as loosely coiled helices extended to varying degrees (1-7, 19, 20). Proteolytic cleavage of the NP protein has been implicated as one of the causes of such variation in nucleocapsid conformation (6, 21); however, in the course of an electron microscopic study of Sendai virus nucleocapsids, we often observed a wide variation in the degree of coiling of duplicate samples that could not be attributed to this mechanism. When ...

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confidence: 73%

“…They are composed of single-stranded RNA and -2600 viral structural protein subunits (NP) with a molecular weight of t60,000 (6,7). Two other proteins (L and P) are associated with the nucleocapsid and thought to be involved in the viral RNA transcriptase activity (8)(9)(10)(11)(12).…”

mentioning

confidence: 99%

Conformation of the helical nucleocapsids of paramyxoviruses and vesicular stomatitis virus: reversible coiling and uncoiling induced by changes in salt concentration.

Heggeness¹,

Scheid²,

Choppin³

1980

Proc. Natl. Acad. Sci. U.S.A.

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“…We note that the C terminus of the protein also interacts, either directly with the capsid, or folds back onto the MoRE as it interacts with the capsid. (5,(28)(29)(30)(31). Nothing is known about the location and conformational state of N TAIL in intact nucleocapsids because N TAIL does not appear to contribute coherently to the reconstructed density from EM, however it is apparent that both the structure and dynamics of the nucleocapsids are significantly modulated by N TAIL .…”

Section: N Tail Populates a Dynamic Equilibrium Comprising Preencodedmentioning

confidence: 99%

Intrinsic disorder in measles virus nucleocapsids

Jensen

Communie

Ribeiro

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

186

217

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The genome of measles virus is encapsidated by multiple copies of the nucleoprotein (N), forming helical nucleocapsids of molecular mass approaching 150 Megadalton. The intrinsically disordered C-terminal domain of N (N TAIL ) is essential for transcription and replication of the virus via interaction with the phosphoprotein P of the viral polymerase complex. The molecular recognition element (MoRE) of N TAIL that binds P is situated 90 amino acids from the folded RNA-binding domain (N CORE ) of N, raising questions about the functional role of this disordered chain. Here we report the first in situ structural characterization of N TAIL in the context of the entire N-RNA capsid. Using nuclear magnetic resonance spectroscopy, small angle scattering, and electron microscopy, we demonstrate that N TAIL is highly flexible in intact nucleocapsids and that the MoRE is in transient interaction with N CORE . We present a model in which the first 50 disordered amino acids of N TAIL are conformationally restricted as the chain escapes to the outside of the nucleocapsid via the interstitial space between successive N CORE helical turns. The model provides a structural framework for understanding the role of N TAIL in the initiation of viral transcription and replication, placing the flexible MoRE close to the viral RNA and, thus, positioning the polymerase complex in its functional environment.is a member of the Paramyxoviridae family of the Mononegavirales order of negative sense, single stranded RNA viruses. The viral genome is encapsidated by multiple copies of the nucleoprotein (N) forming a helical nucleocapsid. Transcription and replication of the viral RNA are initiated by an interaction between N and the polymerase complex, composed of the phosphoprotein (P) and the RNAdependent RNA polymerase (1). N consists of two domains: N CORE (residues 1-400), responsible for the interaction with the viral RNA and for maintaining the nucleocapsid structure, and a long intrinsically disordered domain, N TAIL (residues 401-525) serving as the anchor point for the polymerase complex (2, 3). The molecular recognition element (MoRE) (residues 485-502) of the disordered N TAIL interacts with the C-terminal three-helix bundle domain, XD, of P (residues 459-507) (4) and thereby recruits the polymerase complex onto the nucleocapsid template (5, 6).The realization that intrinsically disordered proteins (IDPs) are functional despite a lack of structure (7-9) has revealed entirely new paradigms that appear to redefine our understanding of the role of conformational flexibility in molecular interactions (10-12). Until now most IDPs have been studied in isolation, or in the presence of a single interaction partner, although it is evident that a real physiological environment could influence the nature and relevance of apparent intrinsic disorder. In this context resolving the question of whether the protein is actually disordered in situ is of paramount importance. In this case the mechanistic role of the extensive disorder present in N TA...

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“…The smooth NC variant is a flexiform linear structure, approximately 1-1 ~tm in length with an 18 nm outer diameter (Finch & Gibbs, 1970;Kingsbury, 1972;Bussell et al, 1974). Well delineated caps id subunits consisting of N protein monomers are arranged in a single-start helical configuration, angled relative to the long axis (Finch, 1970), imparting a herringbone appearance to the NC strands. A second granular NC form has been described for MV and CDV (Koestner & Long, 1970;Robbins et al, 1980;Narang, 1981).…”

Section: Introductionmentioning

confidence: 99%

Isolation and Characterization of Canine Distemper Virus Nucleocapsid Variants

Oglesbee¹,

Tatalick²,

Rice

et al. 1989

Journal of General Virology

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SUMMARYNucleocapsid (NC) variants expressed by the Onderstepoort strain of canine distemper virus (CDV) were ultrastructurally and biochemically characterized. Three isolated variants were defined which corresponded to the three variants observed within the cytoplasm of infected cells. Dense NC (D-NC), isolated on discontinuous caesium chloride (CsCI) isopycnic gradients, had an average density of 1.2971 + 0.0042 g/ml. Ultrastructurally, D-NC were 1620.0 + 112.1 nm in length with a 20.1 _+ 1-3 nm outer and a 5.8 + 0.7 nm inner core diameter. The D-NC protein composition was 89-7 ~ of a 61K protein (N), 8-4 ~ of a 75K protein (P) and 1-9 ~o of a 160K to 200K protein (L). A single species of nucleic acid, 15 kb in length, was isolated from D-NC. Light NC (L-NC), similarly isolated, had an average density of 1-2894+0-0040g/ml. L-NC differed ultrastructurally from D-NC in that poor resolution of NC subunits, a larger outer diameter (32.0 + 2.8 nm), and a greater inner core diameter (10.4 + 0.6 nm) were observed. The average L-NC strand length was 1574.4 + 115.8 nm. The protein composition was the same as D-NC with the exception of an additional 70K protein, representing 4.0 to 7.7 ~ of the total L-NC protein mass. A 15 kb nucleic acid was also identified in L-NC, although heightened sensitivity of encapsidated L-NC nucleic acid to non-specific nuclease degradation was observed. The ratio of D-NC to L-NC isolated from individual virus preparations varied and was independent of viral infectivity. A third NC variant, defective-NC (Df-NC), was also identified. This had the lowest density on CsC1 gradients (1.2460 + 0.0046 g/ml). The Df-NC structures were truncated to a uniform length of 87.0 + 5.8 nm. Diameter measurements were between those of D-NC and L-NC, being 24.4 + 1-4 nm (outer) and 6.9 + 0.4 nm (inner core). Like L-NC, the 70K protein was present but in greater amounts, representing as much as 43-7~ of the total Df-NC protein mass. RNase A-sensitive nucleic acid was isolated from Df-NC which ranged in size from 1.16 to 0-67 kb with a majority of the material being 0.86 kb in length. For both L-NC and Df-NC, canine CDV convalescent serum reacted with viral N and P proteins in Western blot analyses but not with the 70K protein, suggesting a host cellular origin for the latter.

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Observations on the Structure of the Nucleocapsids of some Paramyxoviruses

Cited by 85 publications

References 13 publications

Conformation of the helical nucleocapsids of paramyxoviruses and vesicular stomatitis virus: reversible coiling and uncoiling induced by changes in salt concentration.

Conformation of the helical nucleocapsids of paramyxoviruses and vesicular stomatitis virus: reversible coiling and uncoiling induced by changes in salt concentration.

Intrinsic disorder in measles virus nucleocapsids

Isolation and Characterization of Canine Distemper Virus Nucleocapsid Variants

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