The retinoblastoma protein {pll0 nB) interacts with many cellular proteins in complexes potentially important for its growth-suppressing [unction. We have developed and used an improved version of the yeast two-hybrid system to isolate human cDNAs encoding proteins able to bind pll0 RB. One clone encodes a novel type 1 protein phosphatase catalytic subunit (PP-la2), which differs from the originally defined PP-lc~ by an amino-terminal l 1-amino-acid insert. In vitro-binding assays demonstrated that PP-lc~ isoforms preferentially bind the hypophosphorylated form of p ll0 RB. Moreover, similar pll0 RB sequences are required for binding PP-lc~2 and SV40 large T antigen. Cell cycle synchrony experiments revealed that this association occurs from mitosis to early Gv The implications of these findings on the regulation of both proteins are discussed.
SUMMARY Interactions between tumorigenic cells and their surrounding microenvironment are critical for tumor progression yet remain incompletely understood. Germline mutations in the NF1 tumor suppressor gene cause neurofibromatosis type 1 (NF1), a common genetic disorder characterized by complex tumors called neurofibromas. Genetic studies indicate that biallelic loss of Nf1 is required in the tumorigenic cell of origin in the embryonic Schwann cell lineage. However, in the physiologic state, Schwann cell loss of heterozygosity is not sufficient for neurofibroma formation and Nf1 haploinsufficiency in at least one additional nonneoplastic lineage is required for tumor progression. Here, we establish that Nf1 heterozygosity of bone marrow-derived cells in the tumor microenvironment is sufficient to allow neurofibroma progression in the context of Schwann cell Nf1 deficiency. Further, genetic or pharmacologic attenuation of c-kit signaling in Nf1+/− hematopoietic cells diminishes neurofibroma initiation and progression. Finally, these studies implicate mast cells as critical mediators of tumor initiation.
The retinoblastoma protein interacts with a number of cellular proteins to form complexes which are probably crucial for its normal physiological function. To identify these proteins, we isolated nine distinct clones by direct screening of cDNA expression libraries using purified RB protein as a probe. One of these clones, Apl2, is expressed predominantly at the G1-S boundary and in the S phase of the cell cycle. The nucleotide sequence of Apl2 has features characteristic of transcription factors. The C-terminal region binds to unphosphorylated RB in regions similar to those to which T antigen binds and contains a transactivation domain. A region containing a potential leucine zipper flanked by basic residues is able to bind an E2F recognition sequence specifically. Expression ofApl2 in mammalian cells significantly enhances E2F-dependent transcriptional activity. These results suggest that Apl2 encodes a protein with properties known to be characteristic of transcription factor E2F.The retinoblastoma gene (RB), the first tumor suppressor gene identified, encodes a nuclear phosphoprotein which is ubiquitously expressed in vertebrates (19,20,42,44 (45,47), fluctuates with the cell cycle (7, 11, 13); (ii) the unphosphorylated form of RB is present predominantly in the Go-G1 stage (11, 13); (iii) microinjection of unphosphorylated RB into cells at early G1 inhibits their progression into the S phase (22). These data suggest that RB serves as a critical regulator of entry into the cell cycle and that its inactivation in normal cells leads to deregulated growth.How RB functions is the subject of intense inquiry. Two known biochemical properties of the RB protein have been described; one is its intrinsic DNA-binding activity, which was mapped to its C-terminal 300 amino acid residues (44, 66); another is its ability to interact with several oncoproteins of DNA tumor viruses (12,16,67). This interaction was mapped to two discontinuous regions at amino acids 394 to 571 and 649 to 773, designated as the T-binding domains (29,33). Interestingly, mutations of RB protein in tumors were *
A novel class of visible-light-activated TiO 2 photocatalysts were prepared by direct hydrolysis of tetrabutyl titanate through iodine-doping. When calcination temperature is at 673 K, these nanoparticles (mean diameter of ∼5 nm) show stronger absorption in the 400-550 nm range with a red shift in the band gap transition and significantly higher photocatalytic activity than pure TiO 2 prepared by the same procedure and Degussa P-25 titania nanoparticles in aqueous phenol solution under visible light irradiation (λ > 400 nm). Furthermore, I-doped TiO 2 (673 K) still showed pronounced photocatalytic activity under UV and visible light irradiation.
Phosphatidylglycerol (PG) is an important membrane polyglycerolphospholipid required for the activity of a variety of enzymes and is a precursor for synthesis of cardiolipin and bis(monoacylglycerol) phosphate. PG is subjected to remodeling subsequent to its de novo biosynthesis to incorporate appropriate acyl content for its biological functions and to prevent the harmful effect of lysophosphatidylglycerol (LPG) accumulation. The enzymes involved in the remodeling process have not yet been identified. We report here the identification and characterization of a human gene encoding an acyl-CoA: lysophosphatidylglycerol acyltransferase (LPGAT1). Expression of the LPGAT1 cDNA in Sf9 insect and COS-7 cells led to a significant increase in LPG acyltransferase activity. In contrast, no significant acyltransferase activities were detected against glycerol 3-phosphate or a variety of lysophospholipids, including lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylinositol, and lysophosphatidylserine. The recombinant human LPGAT1 enzyme recognized various acyl-CoAs and LPGs as substrates but demonstrated clear preference to long chain saturated fatty acyl-CoAs and oleoyl-CoA as acyl donors, which is consistent with the lipid composition of endogenous PGs identified from different tissues. Kinetic analyses of LPGAT1 expressed in COS-7 cells showed that oleoyl-LPG was preferred over palmitoyl-LPG as an acyl receptor, whereas oleoylCoA was preferred over lauroyl-CoA as an acyl donor. Consistent with its proposed microsomal origin, LP-GAT1 was localized to the endoplasmic reticulum by subcellular fractionation and immunohistochemical analyses. Northern blot analysis indicated that the human LPGAT1 was widely distributed, suggesting a dynamic functional role of the enzyme in different tissues.
Fanconi anemia (FA) is a chromosomal instability disorder characterized by a progressive bone marrow (BM) failure and an increased incidence of myeloid leukemias. Children with FA are currently being enrolled in clinical trials to evaluate the safety of retroviral-mediated gene transfer. Previously, we used Fancc ؊/؊ mice to show that Fancc ؊/؊ hematopoietic stem cells (HSCs) have a profound defect in repopulating ability. Here, we examined whether retroviral-mediated gene transfer of recombinant Fancc (rFancc) would restore the repopulating ability of Fancc ؊/؊ HSC to wild-type levels. Fancc ؊/؊ HSCs transduced with a retrovirus encoding rFancc exhibited a repopulating ability that approached wild-type levels. Interestingly, ϳ30% of primary recipients (7 of 22) transplanted with uncorrected Fancc ؊/؊ cells developed a range of hematopoietic abnormalities including pancytopenia and BM hypoplasia similar to individuals with FA. Hematopoietic abnormalities were detected in only 1 of 22 mice transplanted with Fancc ؊/؊ cells transduced with a retrovirus encoding rFancc. Moreover, several mice with hematopoietic defects had progenitors that displayed a marked resistance to IFN-␥, TNF-␣, and MIP-1␣ compared to both Fancc ؊/؊ progenitors, which are uniquely hypersensitive to these cytokines, and wild-type progenitors. These data are analogous to studies using progenitors from patients with myelodysplasia and provide functional support for clonal evolution in these mice. Collectively, these data show that gene transfer can enhance HSC repopulating ability and suppresses the tendency for clonal evolution. These studies also reveal potential detrimental effects of ex vivo manipulation for untransduced Fancc ؊/؊
Fanconi anemia (FA) is a heterogenous genetic disease with a high risk of cancer. The FA proteins are essential for interphase DNA damage repair; however, it is incompletely understood why FA-deficient cells also develop gross aneuploidy, leading to cancer. Here, we systematically evaluated the role of the FA proteins in chromosome segregation through functional RNAi screens and analysis of primary cells from patients with FA. We found that FA signaling is essential for the spindle assembly checkpoint and is therefore required for high-fidelity chromosome segregation and prevention of aneuploidy. Furthermore, we discovered that FA proteins differentially localize to key structures of the mitotic apparatus in a cell cycle-dependent manner. The essential role of the FA pathway in mitosis offers a mechanistic explanation for the aneuploidy and malignant transformation known to occur after disruption of FA signaling. Collectively, our findings provide insight into the genetically unstable cancers resulting from inactivation of the FA/BRCA pathway.
A genome-wide screen of phosphatases that control mitosis identified CDKN3, which acts through the CDC2 signaling axis.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.