Reversibly contractile nuclear matrix. Its isolation, structure, and composition.

Abstract

From Tetrahymena macronuclei we have isolated a reversibly contractile nucleoskeleton, i.e., an "expanded" nuclear matrix which reversibly contracts when the total concentration of the bivalent cations, Ca and Mg (3:2), is decreased to 5 mM or increased to 125 raM. During contraction the average diameter of the expanded matrix becomes reduced by about 24%; this corresponds to a volume contraction of about 55%. The reversible contraction of the nuclear matrix does not depend on ATP and cannot be inhibited by salyrgan. The expanded matrix is obtained by removing carefully from the macronuclei 89.7% of the phospholipid, 99,6% of the DNA, 98.5% of the RNA, and 74.8% of the protein by treatment with Triton X-100 and digestion with DNase and RNase followed by an extraction with 2 M NaCl. Electron microscopy reveals, within the expanded matrix, residual equivalents to the structures characteristic for macronuclei: (a) a residual nuclear envelope with nuclear pore complexes; (b) residual nucleoli at the periphery; (c) a fibrillar internal network. The expanded matrix is essentially composed of proteins (96.2%) and traces of DNA (0.8%), RNA (0.5%), phospholipid (1.6%), and carbohydrates (0.9%). The last, which have been determined by gas chromatography, contain glucose, mannose, and an unidentified sugar in the ratio 1:5.4:5.7. The ratio of acidic to basic amino acids of the expanded matrix is 1.55. Sodium dodecyl sulfate (SDS) gel electrophoresis reveals a predominant protein with a mol wt of 18,000 which is apparently involved in the reversible contractile process. The mechanism of this reversible contraction of the expanded matrix remains to be elucidated, but it differs both from actin-myosin contraction systems and from the contractile spasmoneme system in vorticellids.Considerable evidence shows that cell nuclei vary in their size as for example during the cell cycle and cell differentiation, or with circadian periodicity (14). In particular, nuclear enlargement precedes as a prerequisite the onset of DNA synthesis in vivo (12,15) and in vitro (3), while nuclear contraction appears to accompany cessation of DNA synthesis (13). LeStourgeon et aI. (20) have previously suggested that nuclear contractile proteins of the actin-myosin type regulate nuclear contraction and swelling (cf. also reference 9).Recent findings, however, suggest that not actin-myosin type proteins, but rather other acidic proteins determine the internal organization and overall form of cell nuclei. Thus, Berezney and Coffey (4) have isolated, from rat liver nuclei, a T

Isolation and Characterization of Bovine Lymphocyte Nuclear Matrix

“…These results agree with those of nuclear matrices from other sources (2,6,25,30). The significance of these residual nucleic acids is uncertain at present, but there are many reports describing the association of a newly synthesized DNA with nuclear matrices (5,7,9,13,16,27).…”

“…As indicated above, the 68,000 protein may correspond to one of the three major protein components of rat liver nuclear matrix (69,000, 66,000 and 62,000) (6) or nuclear pore complex-lamina (70,000, 66,000 and 62,000) (14). Likewise in rat liver nuclear matrix, three prominent protein bands (60,000-75,000) related to lamin, were usually detected on SDS-gel analysis of nuclear matrix from other sources (9,12,25,30). However, recent studies of the nuclear matrix polypeptides from other cells indicate possible differences in molecular weights.…”

“…The nuclear matrix appears to be a ubiquitous structure that is present in almost all eukaryotic cells. Much evidence is accumulating to suggest that the nuclear matrix not only may function as the skeleton of the nucleus but also may be associated with many dynamic aspects of nuclear function: nuclear swelling (30,31), DNA replication (5,7,13,16,27), RNA transcription (1,11,23), hormone binding (3), T-antigen binding (9), processing and transport of RNA (22,29), etc.…”

“…Since then, some proteinaceous structures, resembling a nuclear matrix, have been prepared from HeLa S3 cell (15), Tetrahymena (30), Friend erythroleukemia cell (19), Physarum polycephalum (25), 3T6 fibroblast (9) and others. These proteinaceous structures are very similar to each other, composed of three components : peripheral lamina, internal fibrogranular structure and residual nucleoli.…”

The major polypeptides of the nuclear pore complex

“…Therefore , such preparations of pore complex-lamina-nuclear matrix remnants from somatic cells are difficult to interpret as to the correlation of the specific structures with the polypeptide components identified by gel electrophoresis. The picture obtained has been further complicated by the finding that somatic nuclei contain a fibrillar network ("nuclear matrix", for synonyms see refs quoted below) that is resistant to extractions with low and high salt buffers , nucleases , and moderate detergents and shows an enrichment of some polypeptides of similar sizes as those found in the nuclear membrane-derived residue fractions mentioned above [8,[15][16][17][18][19][20][21][22][23]. Compared with the somatic nuclei used in the studies mentioned above , the nuclear envelope of the giant nucleus of the amphibian oocyte in mid-to-late stages of oogenesis offers some crucial advantages: (I) It contains a much higher number of pore complexes (ca 4x 1 Q1 per nucleus) in very close packing [3 , 4, 24, 25]…”