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PubMed1995

Transgenic mice expressing an altered murine superoxide dismutase gene provide an animal model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is a progressive neurodegenerative disorder primarily involving motoneurons. A subset of individuals with familial autosomal dominant forms of the disease have mutations of the copper/zinc superoxide dismutase (Cu/Zn SOD, SOD-1) gene, which encodes a ubiquitously expressed enzyme that plays a key role in oxygen free radical scavenging. This observation suggests that altered or reduced SOD-1 activity may play a role in the neurodegenerative process. To explore this possibility further, we have introduced a mutation into the mouse SOD-1 gene that corresponds to one of the changes found in the human gene in familial amyotrophic lateral sclerosis. Integration and expression of this mouse gene in transgenic mice was identified by the presence of a unique restriction enzyme site in the transgene coding sequence generated by introduction of the mutation. We report here that high expression of this altered gene in the central nervous systems of transgenic mice is associated with an age-related rapidly progressive decline of motor function accompanied by degenerative changes of motoneurons within the spinal cord, brain stem, and neocortex. These findings indicate a causative relationship between altered SOD activity and motoneuron degeneration. Moreover, biochemical studies indicate normal levels of total SOD activity in transgenic mouse tissues, results that indicate that the neurodegenerative disorder does not result from a diminution of activity and, as such, represents a dominant "gain of function" mutation.

Age Factors↗
PubMed2004

Caenorhabditis elegans TRPV ion channel regulates 5HT biosynthesis in chemosensory neurons.

Serotonin (5HT) is a pivotal signaling molecule that modulates behavioral and endocrine responses to diverse chemical and physical stimuli. We report cell-specific regulation of 5HT biosynthesis by transient receptor potential V (TRPV) ion channels in C. elegans. Mutations in the TRPV genes osm-9 or ocr-2 dramatically downregulate the expression of the gene encoding the 5HT synthesis enzyme tryptophan hydroxylase (tph-1) in the serotonergic chemosensory neurons ADF, but neither the mutation nor the double mutation of both channel genes affects other types of serotonergic neurons. The TRPV genes are expressed in the ADF neurons but not in other serotonergic neurons, and act cell-autonomously to regulate a neuron-specific transcription program. Whereas in olfactory neurons OSM-9 and OCR-2 function is dependent on ODR-3 Galpha, the activity of ODR-3 or two other Galpha proteins expressed in the ADF neurons is not required for upregulating tph-1 expression, thus the TRPV ion channels in different neurons may be regulated by different mechanisms. A gain-of-function mutation in CaMKII UNC-43 partially suppresses the downregulation of tph-1 in the TRPV mutants, thus CaMKII may be an effector of the TRPV signaling. Mutations in the TRPV genes cause worms developmentally arrest at the Dauer stage. This developmental defect is due in part to reduced 5HT inputs into daf-2/insulin neuroendocrine signaling.

Animals↗
PubMed2002

v-Src SH3-enhanced interaction with focal adhesion kinase at beta 1 integrin-containing invadopodia promotes cell invasion.

In viral Src (v-Src)-transformed cells, focal adhesion kinase (FAK) associates with v-Src by combined v-Src SH2 and gain-of-function v-Src SH3 domain binding to FAK. Here we assess the significance of the Arg-95 to Trp gain-of-function mutation in the v-Src SH3 domain through comparisons of Src-/- fibroblasts transformed with either Prague C v-Src or a point mutant (v-Src-RT) containing a normal (Arg-95) SH3 domain. Both v-Src isoforms exhibited equivalent kinase activity, enhanced Src-/- cell motility, and stimulated cell growth in both low serum and soft agar. The stability of a v-Src-RT.FAK signaling complex and FAK phosphorylation at Tyr-861 and Tyr-925 were reduced in v-Src-RT- compared with v-Src-transformed cells. v-Src but not v-Src-RT promoted Src-/- cell invasion through a reconstituted Matrigel basement membrane barrier and v-Src co-localized with FAK and beta(1) integrin at invadopodia. In contrast, v-Src-RT exhibited a partial perinuclear and focal contact distribution in Src-/- cells. Adenovirus-mediated FAK overexpression promoted v-Src-RT recruitment to invadopodia, the formation of a v-Src-RT.FAK signaling complex, and reversed the v-Src-RT invasion deficit. Adenovirus-mediated inhibition of FAK blocked v-Src-stimulated cell invasion. These studies establish that gain-of-function v-Src SH3 targeting interactions with FAK at beta(1) integrin-containing invadopodia act to stabilize a v-Src.FAK signaling complex promoting cell invasion.

Cell Fusion↗
PubMed2001

Targeting c-kit mutations in solid tumors: scientific rationale and novel therapeutic options.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract. These tumors span a wide clinical spectrum from benign to malignant and have long been recognized for their nearly absolute resistance to chemotherapy and radiation treatment. Surgery is the primary treatment modality for GISTs, but GISTs represent an incurable malignancy for patients with metastatic or unresectable disease. Thus, novel approaches to the treatment of GISTs were desperately needed. Gastrointestinal stromal tumors are characterized by expression of the transmembrane receptor tyrosine kinase KIT, which is defined by the CD117 antigen and is the product of the c-kit proto-oncogene. Activating or gain-of-function mutations in the c-kit gene have been identified in the majority of GIST cases. The resulting constitutive KIT tyrosine kinase activity was hypothesized to provide growth and survival signals to GIST cells and to be crucial to the pathogenesis of the disease. This hypothesis became testable with the identification of the signal transduction inhibitor imatinib mesylate (formerly STI571, [Gleevec]; Novartis Pharmaceuticals Corp, East Hanover, NJ), which blocks the tyrosine kinase activity of KIT as well as the kinase activity of the normal c-abl gene product, the oncogenic Bcr-Abl chimeric fusion protein of chronic myeloid leukemia, and the platelet-derived growth factor receptor. Preclinical experiments showed rapid inhibition of ligand-independent KIT phosphorylation, decreased cellular proliferation, and induction of apoptosis after exposure of GIST cells to imatinib mesylate in vitro. These results provided the rationale to move forward with clinical testing of imatinib mesylate as an anticancer therapy for GIST. In early 2000, a dramatic clinical and radiographic response to imatinib mesylate was shown in a single patient with advanced, chemotherapy-resistant GIST. The powerful scientific rationale for this proof-of-concept study, together with the durable and significant response observed in this first GIST patient treated with imatinib mesylate, have provided the driving force for rapid clinical development of this targeted therapy in this solid tumor indication.

Antineoplastic Agents↗
PubMed2003

Gap junction protein beta 1 (GJB1) mutations and central nervous system symptoms in X-linked Charcot-Marie-Tooth disease.

OBJECTIVES: To clarify the clinical variability, including central nervous system (CNS) involvement, in X-linked Charcot-Marie-Tooth disease (CMTX) patients. MATERIAL AND METHODS: We clinically, pathologically and genetically studied six CMTX patients with distinct symptoms and four different GJB1 mutations. RESULTS: One patient with Val63Ile had deafness, low intelligence, saccadic eye movement, upper extremity distal dominant muscle weakness and normal sensation. Another patient with Glu186Lys had severe sensorineural deafness at the age of 6 years, but did not develop muscle weakness until the age of 20 years. Two patients with Arg22Gln had typical CMT1A-like clinical features, no CNS symptoms and obvious onion bulb formations. Two siblings with deletion of the entire GJB1 gene had mild to moderate lower extremity muscle weakness and sensory disturbance without CNS involvement. CONCLUSION: These findings suggest that some gain of function mutations of GJB1 may be related to CNS symptoms because the patients with GJB1 deletion only had peripheral neuropathy, although other unknown associated factors may contribute to their clinical phenotypes.

Adult↗
PubMed2004

In silico study of action potential and QT interval shortening due to loss of inactivation of the cardiac rapid delayed rectifier potassium current.

The rapid delayed rectifier K(+) current, I(Kr), plays a key role in repolarisation of cardiac ventricular action potentials (APs). In recent years, a novel clinical condition denoted the short QT syndrome (SQTS) has been identified and, very recently, gain in function mutations in the gene encoding the pore-forming sub-unit of the I(Kr) channel have been proposed to underlie SQTS in some patients. Here, computer simulations were used to investigate the effects of the selective loss of voltage-dependent inactivation of I(Kr) upon ventricular APs and on the QT interval of the electrocardiogram. I(Kr) and inactivation-deficient I(Kr) were incorporated into Luo-Rudy ventricular AP models. Inactivation-deficient I(Kr) produced AP shortening that was heterogeneous between endocardial, mid-myocardial, and epicardial ventricular cell models, irrespective of whether heterogeneity between these sub-regions was incorporated of slow delayed rectifier K(+) current (I(Ks)) alone, or of I(Ks) together with that of transient outward K(+) current. The selective loss of rectification of I(Kr) did not augment transmural dispersion of AP repolarisation, as AP shortening was greater in mid-myocardial than in endo- or epicardial cell models. Simulated conduction through a 1 D transmural ventricular strand was altered by incorporation of inactivation-deficient I(Kr) and the reconstructed QT interval was shortened. Collectively, these results substantiate the notion that selective loss of I(Kr) inactivation produces a gain in I(Kr) function that causes QT interval shortening.

Action Potentials↗
PubMed2000

Mutation of the gene encoding the ROR2 tyrosine kinase causes autosomal recessive Robinow syndrome.

Robinow syndrome is a short-limbed dwarfism characterized by abnormal morphogenesis of the face and external genitalia, and vertebral segmentation. The recessive form of Robinow syndrome (RRS; OMIM 268310), particularly frequent in Turkey, has a high incidence of abnormalities of the vertebral column such as hemivertebrae and rib fusions, which is not seen in the dominant form. Some patients have cardiac malformations or facial clefting. We have mapped a gene for RRS to 9q21-q23 in 11 families. Haplotype sharing was observed between three families from Turkey, which localized the gene to a 4. 9-cM interval. The gene ROR2, which encodes an orphan membrane-bound tyrosine kinase, maps to this region. Heterozygous (presumed gain of function) mutations in ROR2 were previously shown to cause dominant brachydactyly type B (BDB; ref. 7). In contrast, Ror2-/- mice have a short-limbed phenotype that is more reminiscent of the mesomelic shortening observed in RRS. We detected several homozygous ROR2 mutations in our cohort of RRS patients that are located upstream from those previously found in BDB. The ROR2 mutations present in RRS result in premature stop codons and predict nonfunctional proteins.

Abnormalities, Multiple↗
PubMed1999

A mutation in the extracellular cysteine-rich repeat region of the beta3 subunit activates integrins alphaIIbbeta3 and alphaVbeta3.

Inside-out signaling regulates the ligand-binding function of integrins through changes in receptor affinity and/or avidity. For example, alphaIIbbeta3 is in a low-affinity/avidity state in resting platelets, and activation of the receptor by platelet agonists enables fibrinogen to bind. In addition, certain mutations and truncations of the integrin cytoplasmic tails are associated with a high-affinity/avidity receptor. To further evaluate the structural basis of integrin activation, stable Chinese hamster ovary (CHO) cell transfectants were screened for high-affinity/avidity variants of alphaIIbbeta3. One clone (AM-1) expressed constitutively active alphaIIbbeta3, as evidenced by (1) binding of soluble fibrinogen and PAC1, a ligand-mimetic antialphaIIbbeta3 antibody; and (2) fibrinogen-dependent cell aggregation. Sequence analysis and mutant expression in 293 cells proved that a single amino acid substitution in the cysteine-rich, extracellular portion of beta3(T562N) was responsible for receptor activation. In fact, T562N also activated alphaVbeta3, leading to spontaneous binding of soluble fibrinogen to 293 cells. In contrast, neither T562A nor T562Q activated alphaIIbbeta3, suggesting that acquisition of asparagine at residue 562 was the relevant variable. T562N also led to aberrant glycosylation of beta3, but this was not responsible for the receptor activation. The binding of soluble fibrinogen to alphaIIbbeta3(T562N) was not sufficient to trigger tyrosine phosphorylation of pp125(FAK), indicating that additional post-ligand binding events are required to activate this protein tyrosine kinase during integrin signaling. These studies have uncovered a novel gain-of-function mutation in a region of beta3 intermediate between the ligand-binding region and the cytoplasmic tail, and they suggest that this region is involved in integrin structural changes during inside-out signaling.

Amino Acid Substitution↗
PubMed2000

Knots in the family tree: evolutionary relationships and functions of knox homeobox genes.

Knotted-like homeobox (knox) genes constitute a gene family in plants. Class I knox genes are expressed in shoot apical meristems, and (with notable exceptions) not in lateral organ primordia. Class II genes have more diverse expression patterns. Loss and gain of function mutations indicate that knox genes are important regulators of meristem function. Gene duplication has contributed to the evolution of families of homeodomain proteins in metazoans. We believe that similar mechanisms have contributed to the diversity of knox gene function in plants. Knox genes may have contributed to the evolution of compound leaves in tomato and could be involved in the evolution of morphological traits in other species. Alterations in cis-regulatory regions in some knox genes correlate with novel patterns of gene expression and distinctive morphologies. Preliminary data from the analysis of class I knox gene expression illustrates the evolution of complex patterns of knox expression is likely to have occurred through loss and gain of domains of gene expression.

Evolution, Molecular↗
PubMed2026

The genomic origins and evolutionary path to a key innovation in the world's most venomous snakes.

Evolutionary innovation is a catalyst for the colonization of new environments and the adaptive radiations of major groups. Novel traits typically evolve through the modification of preexisting characters, but the genetic paths underlying their origin have been challenging to trace, and the general requirements for and relative order of different kinds of gene mutations have been difficult to assess. Here, we trace the genomic origins of four procoagulant venom toxins (factor X, factor V, group I phospholipase A2, and Kunitz-type toxins) that collectively underlie a novel, especially potent blood-clotting venom type in the recently evolved Australian brown snake and taipan clade. We find evidence for a previously unknown fifth toxin, coagulation factor VII, and show that the toxins evolved through two distinct genetic paths. The factor X and factor V toxins evolved through the sequential de novo co-option of ancestral clotting factor proteins that entailed their heterotopic expression in the venom gland, the fixation of segmental duplications containing each locus, and subsequent gain-of-function mutations that rendered factor X and factor V constitutively active. In contrast, the phospholipase A2 and Kunitz-type toxins evolved by modifying the functions of neurotoxins that were part of the venom arsenal. Our findings support models in which innovative mutations in single-copy genes precede gene duplication in the evolution of novel proteins and offer a rare view into the genesis of a complex trait that has played a central role in a major adaptive radiation.

Animals↗
PubMed2026

The genomic origins and evolutionary path to a key innovation in the world's most venomous snakes.

Evolutionary innovation is a key driver of the colonization of new environments and the adaptive radiations of major groups. Novel traits typically evolve through the modification of pre-existing characters but the genetic paths underlying their origin have been challenging to trace, and the general requirements for and relative order of different kinds of gene mutations have been difficult to assess. Here, we trace the genomic origins of four procoagulant venom toxins (factor X, factor V, group I phospholipase A2, and Kunitz-type toxins) that collectively underlie a novel, especially potent blood-clotting venom type in the recently evolved Australian brown snake and taipan clade. We discover evidence for a previously unknown fifth toxin, coagulation factor VII, and show that the toxins evolved through two distinct genetic paths. The factor X and factor V toxins evolved through the sequential de novo co-option of ancestral clotting factor proteins that entailed their heterotopic expression in the venom gland, the fixation of segmental duplications containing each locus, and subsequent gain-of-function mutations that rendered factor X and factor V constitutively active. In contrast, the phospholipase A2 and Kunitz-type toxins evolved by modifying the functions of neurotoxins that were part of the venom arsenal. Our findings support models in which innovative mutations in single-copy genes precede gene duplication in the evolution of novel proteins and offer a rare view into the genesis of a complex trait that has played a central role in a major adaptive radiation.

Biological Sciences: Evolution↗
PubMed1998

Yeast gene YRR1, which is required for resistance to 4-nitroquinoline N-oxide, mediates transcriptional activation of the multidrug resistance transporter gene SNQ2.

We have cloned and characterized a Saccharomyces cerevisiae gene YRR1 that is important for resistance to the mutagen 4-nitroquinoline N-oxide (4-NQO). The wild-type YRR1 gene encodes a protein that contains a Zn(II)2Cys6-type zinc-finger motif. Disruption of the YRR1 gene leads to hypersensitivity to 4-NQO. A dominant mutation (YRR1-1) that confers strong resistance to 4-NQO has been identified. Epistasis analysis demonstrated that 4-NQO resistances mediated by the YRR1 and YRR1-1 alleles require the presence of the SNQ2 gene that encodes a multidrug resistance ATP binding cassette superfamily protein responsible for 4-NQO export. Northern blot analysis of SNQ2 mRNA levels indicated that Yrr1p is involved in basal and drug-induced transcriptional activation of SNQ2, whereas Pdr1p/Pdr3p transcription factors are mainly involved in basal SNQ2 expression. In the YRR1-1 mutant, the level of SNQ2 mRNA is constitutively elevated. These results establish that Yrr1p is important for 4-NQO resistance by mediating transcriptional activation of the SNQ2 gene in response to the stress imposed by 4-NQO. The gain-of-function mutation of Yrr1-1p was attributable to the duplication of a 12-amino-acid sequence generated near the carboxy terminus.

4-Nitroquinoline-1-oxide↗
PubMed2002

Organ-specific cell division abnormalities caused by mutation in a general cell cycle regulator in C. elegans.

The precise control of cell division during development is pivotal for morphogenesis and the correct formation of tissues and organs. One important gene family involved in such control is the p21/p27/p57 class of negative cell cycle regulators. Loss of function of the C. elegans p27 homolog, cki-1, causes extra cell divisions in numerous tissues including the hypodermis, the vulva, and the intestine. We have sought to better understand how cell divisions are controlled upstream or in parallel to cki-1 in specific organs during C. elegans development. By taking advantage of the invariant cell lineage of C. elegans, we used an intestinal-specific GFP reporter in a screen to identify mutants that undergo cell division abnormalities in the intestinal lineage. We have isolated a mutant with twice the wild-type complement of intestinal cells, all of which arise during mid-embryogenesis. This mutant, called rr31, is a fully dominant, maternal-effect, gain-of-function mutation in the cdc-25.1 cell cycle phosphatase that sensitizes the intestinal lineage to an extra cell division. We showed that cdc-25.1 acts at the G1/S transition, as ectopic expression of CDC-25.1 caused entry into S phase in intestinal cells. In addition, we showed that the cdc-25.1(gf) requires cyclin E. The extra cell division defect was shown to be restricted to the E lineage and the E fate is necessary and sufficient to sensitize cells to this mutation.

Amino Acid Sequence↗
PubMed1998

Somatic mutations that contribute to breast cancer.

Cytogenetic and molecular analyses of primary sporadic human breast carcinomas have documented at least 12 different chromosome arms affected by loss of heterozygosity (LOH). This has been taken as evidence for the presence of putative tumour suppressor genes in the remaining allele within the affected regions. We have previously identified three regions on chromosome 17q that are affected by LOH in primary human breast tumours. A physical map of one of these regions (17q21) has been prepared. The putative target gene appears to be located between the D17S846 and D17S746 loci. We are currently determining whether either of two genes located in this region is the target for LOH. The mouse mammary tumour model system provides an approach for identifying genes which, when activated or inactivated by mouse mammary tumour virus (MMTV) integration, contribute to specific stages of mammary tumorigenesis. Using this approach we have identified two genes, designated NOTCH4/INT3 and INT6 respectively. Interruption of NOTCH4/INT3 by MMTV represents a gain-of-function mutation that has profound consequences for mammary gland development and tumorigenesis. INT6 was found to be interrupted by an integrated MMTV genome in a mammary hyperplastic outgrowth line and two independent mammary tumours. In each case the transcriptional orientation of the viral genome was opposite to that of INT6. The rearranged allele was expressed as a truncated chimaeric RNA species composed of INT6 coding sequences, intron sequences and MMTV sequences. Since the non-rearranged allele contained no mutations, we conclude that MMTV integration into INT6 causes a dominant-negative mutation or biologically activates its function. The nucleotide sequence of INT6 is unrelated to any of the known genes in the GenBank database, but is evolutionarily highly conserved.

Animals↗
PubMed2004

Mild overexpression of MeCP2 causes a progressive neurological disorder in mice.

Mutations in the X-linked methyl-CpG-binding protein 2 (MECP2), encoding a transcriptional repressor, cause Rett syndrome and a variety of related neurodevelopmental disorders. The vast majority of mutations associated with human disease are loss-of-function mutations, but precisely what aspect of MeCP2 function is responsible for these phenotypes remains unknown. We overexpressed wild-type human protein in transgenic mice using a large genomic clone containing the entire human MECP2 locus. Detailed neurobehavioral and electrophysiological studies in transgenic line MeCP2(Tg1), which expresses MeCP2 at approximately 2-fold wild-type levels, demonstrated onset of phenotypes around 10 weeks of age. Surprisingly, these mice displayed enhanced motor and contextual learning and enhanced synaptic plasticity in the hippocampus. After 20 weeks of age, however, these mice developed seizures, became hypoactive and approximately 30% of them died by 1 year of age. These data demonstrate that MeCP2 levels must be tightly regulated in vivo, and that even mild overexpression of this protein is detrimental. Furthermore, these results support the possibility that duplications or gain-of-function mutations in MECP2 might underlie some cases of X-linked delayed-onset neurobehavioral disorders.

Animals↗
PubMed2001

BIN2, a new brassinosteroid-insensitive locus in Arabidopsis.

Brassinosteroids (BRs) play important roles throughout plant development. Although many genes have been identified that are involved in BR biosynthesis, genetic approaches in Arabidopsis have led to the identification of only one gene, BRI1, that encodes a membrane receptor for BRs. To expand our knowledge of the molecular mechanism(s) of plant steroid signaling, we analyzed many dwarf and semidwarf mutants collected from our previous genetic screens and identified a semidwarf mutant that showed little response to exogenous BR treatments. Genetic analysis of the bin2 (BR-INSENSITIVE 2) mutant indicated that the BR-insensitive dwarf phenotype was due to a semidominant mutation in the BIN2 gene that mapped to the middle of chromosome IV between the markers CH42 and AG. A direct screening for similar semidwarf mutants resulted in the identification of a second allele of the BIN2 gene. Despite some novel phenotypes observed with the bin2/+ mutants, the homozygous bin2 mutants were almost identical to the well-characterized bri1 mutants that are defective in BR perception. In addition to the BR-insensitive dwarf phenotype, bin2 mutants exhibited BR insensitivity when assayed for root growth inhibition and feedback inhibition of CPD gene expression. Furthermore, bin2 mutants displayed an abscisic acid-hypersensitive phenotype that is shared by the bri1 and BR-deficient mutants. A gene dosage experiment using triploid plants suggested that the bin2 phenotypes were likely caused by either neomorphic or hypermorphic gain-of-function mutations in the BIN2 gene. Thus, the two bin2 mutations define a novel genetic locus whose gene product might play a role in BR signaling.

Arabidopsis↗
PubMed2004

Mouse model of Noonan syndrome reveals cell type- and gene dosage-dependent effects of Ptpn11 mutation.

Noonan syndrome is a common human autosomal dominant birth defect, characterized by short stature, facial abnormalities, heart defects and possibly increased risk of leukemia. Mutations of Ptpn11 (also known as Shp2), which encodes the protein-tyrosine phosphatase Shp2, occur in approximately 50% of individuals with Noonan syndrome, but their molecular, cellular and developmental effects, and the relationship between Noonan syndrome and leukemia, are unclear. We generated mice expressing the Noonan syndrome-associated mutant D61G. When homozygous, the D61G mutant is embryonic lethal, whereas heterozygotes have decreased viability. Surviving Ptpn11(D61G/+) embryos ( approximately 50%) have short stature, craniofacial abnormalities similar to those in Noonan syndrome, and myeloproliferative disease. Severely affected Ptpn11(D61G/+) embryos ( approximately 50%) have multiple cardiac defects similar to those in mice lacking the Ras-GAP protein neurofibromin. Their endocardial cushions have increased Erk activation, but Erk hyperactivation is cell and pathway specific. Our results clarify the relationship between Noonan syndrome and leukemia and show that a single Ptpn11 gain-of-function mutation evokes all major features of Noonan syndrome by acting on multiple developmental lineages in a gene dosage-dependent and pathway-selective manner.

Animals↗
PubMed1996

The mitogen-activated protein kinase phosphatases PAC1, MKP-1, and MKP-2 have unique substrate specificities and reduced activity in vivo toward the ERK2 sevenmaker mutation.

Mitogen-activated protein (MAP) kinases can be grouped into three structural families, ERK, JNK, and p38, which are thought to carry out unique functions within cells. We demonstrate that ERK, JNK, and p38 are activated by distinct combinations of stimuli in T cells that simulate full or partial activation through the T cell receptor. These kinases are regulated by reversible phosphorylation on Tyr and Thr, and the dual specific phosphatases PAC1 and MKP-1 previously have been implicated in the in vivo inactivation of ERK or of ERK and JNK, respectively. Here we characterize a new MAP kinase phosphatase, MKP-2, that is induced in human peripheral blood T cells with phorbol 12-myristate 13-acetate and is expressed in a variety of nonhematopoietic tissues as well. We show that the in vivo substrate specificities of individual phosphatases are unique. PAC1, MKP-2, and MKP-1 recognize ERK and p38, ERK and JNK, and ERK, p38, and JNK, respectively. Thus, individual MAP kinase phosphatases can differentially regulate the potential for cross-talk between the various MAP kinase pathways. A hyperactive allele of ERK2 (D319N), analogous to the Drosophila sevenmaker gain-of-function mutation, has significantly reduced sensitivity to all three MAP kinase phosphatases in vivo.

Animals↗