Topoisomerase II is required for mitoxantrone to signal NFkB activation in HL60 cells

Boland, M.P.

doi:10.1074/jbc.m003794200

Cited by 6 publications

(2 citation statements)

References 45 publications

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“…The presence of topoisomerase IIα (TOP IIα) in the complex is indicative of a possible dual role for the CDC5L complex in the cell nucleus. It has been suggested that TOP IIα may be involved in transcription because of its enhanced expression during mitosis (Boland et al ., 2000).…”

Section: Discussionmentioning

confidence: 99%

Functional analysis of the human CDC5L complex and identification of its components by mass spectrometry

Küster²,

et al. 2000

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Recently, we identi®ed proteins that co-purify with the human spliceosome using mass spectrometry. One of the identi®ed proteins, CDC5L, corresponds to the human homologue of the Schizosaccharomyces pombe CDC5 + gene product. Here we show that CDC5L is part of a larger multiprotein complex in HeLa nuclear extract that incorporates into the spliceosome in an ATP-dependent step. We also show that this complex is required for the second catalytic step of pre-mRNA splicing. Immunodepletion of the CDC5L complex from HeLa nuclear extract inhibits the formation of pre-mRNA splicing products in vitro but does not prevent spliceosome assembly. The ®rst catalytic step of pre-mRNA splicing is less affected by immunodepleting the complex. The puri®ed CDC5L complex in HeLa nuclear extract restores pre-mRNA splicing activity when added to extracts that have been immunodepleted using anti-CDC5L antibodies. Using mass spectrometry and database searches, the major protein components of the CDC5L complex have been identi®ed. This work reports a ®rst puri®cation and characterization of a functional, human non-snRNA spliceosome subunit containing CDC5L and at least ®ve additional protein factors. Keywords: CDC5L/mass spectrometry/pre-mRNA splicing/spliceosome IntroductionNuclear pre-mRNA splicing is the process by which the introns (intervening sequences) in the primary transcript are speci®cally removed and the coding sequences joined to form mature mRNA, which is subsequently transported into the cytoplasm for protein synthesis. Splicing takes place in the nucleus, via a two-step transesteri®cation mechanism, and this is catalysed by a large RNA±protein complex termed the spliceosome. The major subunits of spliceosomes are the U1, U2, U5 and U4/U6 small nuclear ribonucleoprotein particles (snRNPs), each of which contains the corresponding snRNA and a set of snRNP proteins (reviewed by Kra Èmer, 1996;Will and Lu Èrhmann, 1997). In addition, other non-snRNP protein splicing factors are also required for spliceosome formation and splicing (Will and Lu Èrhmann, 1997;Staley and Guthrie, 1998). Spliceosome assembly occurs in a pathway that involves the sequential binding of snRNP and protein splicing factors to conserved intron sequences to form the active complex. This is a multistep process and separate ATP-dependent complexes designated A, B and C have been identi®ed in vitro (reviewed by Reed and Palandjian, 1997). The C complex contains the active spliceosome while the A and B complexes contain assembly intermediates. The detailed roles played by many spliceosomal proteins in the pre-mRNA splicing mechanism are not yet fully understood.The protein composition of mammalian spliceosomes has been studied using complexes puri®ed from HeLa nuclear extracts by a combination of gel ®ltration and af®nity chromatography (Reed, 1990;Bennett et al., 1992). More recently in our laboratories, a large-scale analysis of spliceosome-associated proteins has been carried out using mass spectrometry and database searches (Neubauer et al., 1998). One...

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Section: Discussionmentioning

confidence: 99%

Functional analysis of the human CDC5L complex and identification of its components by mass spectrometry

Küster²,

et al. 2000

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“…A possible mechanism for therapeutic efficacy of mitoxantrone in treatment of multiple sclerosis is that it may inhibit migration of inflammatory cells into and within the central nervous system. DNA damage resulting due to poisoning of topoisomerase II by mitoxantrone acts as a signal for NF-kappa B activation and induction of apoptosis (Boland et al 2000). Mitoxantrone is 78 % bound to plasma proteins (Scott and Figgitt 2004;Wiseman and Spencer 1997).…”

Section: Mitoxantronementioning

confidence: 99%

Multi-Targeted Approach to Treatment of Cancer

Gandhi¹,

Mehta²,

Grover³

et al. 2015

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Roles of DNA topoisomerase II isozymes in chemotherapy and secondary malignancies

Azarova¹,

Lyu²,

Lin³

et al. 2007

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

262

222

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Drugs that target DNA topoisomerase II (Top2), including etoposide (VP-16), doxorubicin, and mitoxantrone, are among the most effective anticancer drugs in clinical use. However, Top2-based chemotherapy has been associated with higher incidences of secondary malignancies, notably the development of acute myeloid leukemia in VP-16-treated patients. This association is suggestive of a link between carcinogenesis and Top2-mediated DNA damage. We show here that VP-16-induced carcinogenesis involves mainly the ␤ rather than the ␣ isozyme of Top2. In a mouse skin carcinogenesis model, the incidence of VP-16-induced melanomas in the skin of 7,12-dimethylbenz[a]anthracene-treated mice is found to be significantly higher in TOP2␤ ؉ than in skin-specific top2␤-knockout mice. Furthermore, VP-16-induced DNA sequence rearrangements and double-strand breaks (DSBs) are found to be Top2␤-dependent and preventable by cotreatment with a proteasome inhibitor, suggesting the importance of proteasomal degradation of the Top2␤-DNA cleavage complexes in VP-16-induced DNA sequence rearrangements. VP-16 cytotoxicity in transformed cells expressing both Top2 isozymes is, however, found to be primarily Top2␣-dependent. These results point to the importance of developing Top2␣-specific anticancer drugs for effective chemotherapy without the development of treatment-related secondary malignancies.DNA rearrangements ͉ melanoma ͉ skin-specific topoisomerase II␤-knockout ͉ tumor cell killing ͉ carcinogenesis A nticancer drugs that target DNA topoisomerase II (Top2), including etoposide (VP-16), doxorubicin, and mitoxantrone, are often referred to as Top2 poisons and are among the most effective and widely used anticancer drugs in the clinic. However, life-threatening toxic side effects, including drug-induced secondary malignancies, have been noted in patients receiving Top2-based chemotherapy. An association between infant leukemia and in utero exposure to Top2 poisons has also been reported (reviewed in refs. 1-3). In all cases, the molecular basis underlying carcinogenesis in Top2-based chemotherapy is unclear.Clinical evidence for a direct link between VP-16 treatment and treatment-related acute myeloid leukemia (t-AML) is particularly strong (1-3). VP-16-induced t-AML is frequently associated with balanced translocations between the mixed lineage leukemia (MLL) gene on chromosome 11q23 and Ͼ50 partner genes (the MLL gene is also known as ALL-1, hTRX, or HRX) (4-7). These rearrangements, as well as those found in infant leukemia, cluster within a well characterized 8.3-kb breakpoint cluster region (bcr) (8-16). The bcr of MLL is AT-rich and contains Alu sequences, putative recognition sites of Top2-mediated DNA cleavage, and chromosome scaffold/matrix attachment regions (SAR/MAR) (5,(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). There is substantial evidence that chromosome 11q23 translocations in t-AML and infant leukemia are a consequence of drug-induced formation of double-strand breaks (DSBs) (6-9). VP-16 is known to induce DSBs by the format...

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Topoisomerase II is required for mitoxantrone to signal NFkB activation in HL60 cells

Cited by 6 publications

References 45 publications

Functional analysis of the human CDC5L complex and identification of its components by mass spectrometry

Functional analysis of the human CDC5L complex and identification of its components by mass spectrometry

Multi-Targeted Approach to Treatment of Cancer

Roles of DNA topoisomerase II isozymes in chemotherapy and secondary malignancies

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