Association of Lyn Tyrosine Kinase with the Nuclear Matrix and Cell‐Cycle‐Dependent Changes in Matrix‐Associated Tyrosine Kinase Activity

Abstract

The nuclear matrix isolated from HeLa cells and Rat2 fibroblasts harbor? tyrosine kinase and tyrosine phosphatase activities. Polypeptides of 53, 56 and 60 kDa, associated with this subnuclear structure, were phosphorylated at tyrosine in vivo. By immunoblot and immunolabelling experiments, we identified one of the nuclear-matrix-associated tyrosine kinases as Lyn, a Src family member. Lyn was distributed as foci throughout the matrix. The p56 and p53 isoforms of Lyn remained firmly associated with the nuclear matrix after a variety of matrix preparation procedures, and were not detectable in the chromatin fraction of the nucleus. The tyrosine kinase activity associated with the nuclear matrix showed cell-cycle-dependent changes, maximum activity being observed at the G,/S transition phase. Polyoma-virus-transformed rat fibroblast cells showed sixfold higher tyrosine kinase activity in the nuclear matrix preparations compared to that in untransformed cells. These observations are consistent with the suggestion that tyrosine kinase activity associated with the nuclear matrix may be an important determinant of cellular proliferation.Keywords: nuclear matrix ; Lyn tyrosine kinase ; cell cycle ; protein phosphorylation ; nuclear tyrosine kinase.Phosphorylation at tyrosine residues regulates properties of proteins such as enzyme activity, subcellular localization and intermolecular interaction. In eukaryotic cells, this modification effects the response of the cell to its external environment by helping to transmit signals from the cell membrane across the cytoplasm to the nucleus, where actual control of gene expression takes place [I]. The intracellular protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs) are either associated with various structures such as the plasma membrane, endoplasmic reticulum, cytoskeleton, spindle fibres and mitochondria or are present in a soluble form in the cytoplasm [2, 31. The activities and substrate specificities of these enzymes in vivo are determined by phosphorylation, association with other proteins and subcellular compartmentalization [4, 51.Recent studies have identified the presence of tyrosine-phosphorylated proteins in the nucleus and their role in cell-cycle control and as transcription factors [6, 71. Both cytoplasmic and nuclear proteins are phosphorylated at tyrosine residues in the signalling pathway of mitogenesis. A few tyrosine kinases and phosphatases have been shown to localize to the nucleus, but their physiological functions have not been well defined 13, 81. PTKs known to localize to the nucleus constitutively are Wee 1, Abl, Fer/Fer T and Fgr 181. The Src tyrosine kinase, which is the prototype member of the Src family of kinases, is an ubiqui- tously expressed cytoplasmic enzyme and has been shown to be localized in the nucleus of calcium-induced differentiating keratinocytes 191. Studies on the subnuclear distribution of the tyrosine phosphorylated proteins and the PTKs and PTPs are generally lacking and could provide clues to the role of...

Two Splice Variants of a Tyrosine Phosphatase Differ in Substrate Specificity, DNA Binding, and Subcellular Location

“…(33). Recently tyrosine kinase activity has been found in the nuclear matrix, which shows cell cycle-dependent changes that are maximal during G 1 /S transition (27). One of the tyrosine kinases found tightly associated with nuclear matrix is Lyn, a member of the Src family (27).…”

“…Nuclear Matrix Preparation and Binding of PTP-S2, PTP-S4, and ⌬PTP-S4 -Nuclear matrix was prepared as described previously (26,27). The nuclear matrix preparation was suspended in buffer A (10 mM HEPES, pH 8.0, 5 mM KCl, 3 mM MgCl 2 , 1 mM phenylmethylsulfonyl fluoride, 2 g/ml soybean trypsin inhibitor, 2 g/ml leupeptin and aprotinin) containing 150 mM NaCl, and incubated on ice for 30 min with various amounts of either PTP-S2, PTP-S4, or ⌬PTP-S4 (or control buffer).…”

Lapatinib Antagonizes Multidrug Resistance-Associated Protein 1-Mediated Multidrug Resistance by Inhibiting Its Transport Function

“…Strict control of the tyrosine kinase activity in the cell regulates important processes such as cell cycle, proliferation and apoptosis (44,45 (46,47). Other TKIs including gefitinib, erlotinib and vande- tanib have shown the ability to antagonize the function of ABCB1 and ABCG2 (36,48,49).…”

Role of transforming growth factor and vascular endothelial growth factor and their receptors in the pathogenesis of bleomycin induced lung fibrosis

“…This in turn activates the SIRT1 transcription and suppresses the transcription of MMP-9 promoter by deacetylation of NF-kB eventually resulting in chronic progressive interstitial pulmonary fibrosis Role of receptor tyrosine kinases (RTKs) in pulmonary fibrosis The tyrosine kinases are a sub-class of protein kinase enzyme that can transfer a phosphate group from ATP to cellular proteins. Phosphorylation at tyrosine residues controls many cellular functions by regulating enzyme activity, subcellular localization, and interaction between molecules and growth factors leading to downstream cell signaling (Radha et al, 1996, Schaller et al, 1992. These cell signal transduction cascades transmit the extracellular signals through the cell membrane to the cytoplasm and to the nucleus, leading to altered gene expression (Radha et al, 1996) and ultimately altered cellular functions.…”

“…Phosphorylation at tyrosine residues controls many cellular functions by regulating enzyme activity, subcellular localization, and interaction between molecules and growth factors leading to downstream cell signaling (Radha et al, 1996, Schaller et al, 1992. These cell signal transduction cascades transmit the extracellular signals through the cell membrane to the cytoplasm and to the nucleus, leading to altered gene expression (Radha et al, 1996) and ultimately altered cellular functions. If the tyrosine kinases become constitutively active, a non-stop functional state develops that may contribute to initiation or progression of cancer.…”