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PubMed1997

Hb Osler [beta 145(HC2)Tyr-->Asp] results from posttranslational modification.

We studied two members of an African American family with erythrocytosis. An abnormal hemoglobin variant with an electrophoretic pattern on cellulose acetate similar to Hb J was identified. The oxygen dissociation curve using whole blood was biphasic, dramatically left-shifted, and hyperbolic. Sequence analysis of DNA from the proband showed heterozygosity for a T-->A change at the first position of codon 145 in the beta-globin gene which results in the substitution of an asparagine residue for normal tyrosine. The second cycle of C-terminal amino acid sequence analysis of a mixture of alpha- and beta-globin chains showed tyrosine, aspartic acid, and small amounts of asparagine. Collectively, these results indicate the existence of a mutation at codon 145 of the beta-globin gene which encodes for asparagine instead of tyrosine, and that asparagine then undergoes a partial posttranslational deamidation to aspartic acid. This amino acid substitution corresponds to Hb Osler, which is a high oxygen affinity hemoglobin variant, initially described to be caused by a substitution of Tyr-->Asp at beta 145. Posttranslational amino acid modification may constitute an important component in the pathophysiology of hemoglobinopathies.

Adult↗

PubMed1988

Posttranslational modifications of alpha tubulin: detyrosination and acetylation differentiate populations of interphase microtubules in cultured cells.

Subsets of microtubules enriched in posttranslationally detyrosinated (Gundersen, G. G., M. H. Kalnoski, and J. C. Bulinski. 1984. Cell. 38:779) or acetylated (Piperno, G., M. Le Dizet, and X. Chang. 1987. J. Cell Biol. 104:298), alpha tubulin have previously been described in interphase cultured cells. In this study an immunofluorescence comparison of these minor populations of microtubules revealed that, in African green monkey kidney epithelial cells (TC-7 line), the population of microtubules enriched in detyrosinated tubulin was virtually coincident with the population enriched in acetylated alpha tubulin. In some cell types, however, such as human HeLa or marsupial PtK-2 cells, only one posttranslationally modified form of tubulin, i.e., acetylated or detyrosinated, respectively, was detectable in microtubules. In TC-7 cells, although both modifications were present, dissimilar patterns and kinetics of reappearance of microtubules enriched in detyrosinated and acetylated tubulin were observed after recovery of cells from microtubule-depolymerizing treatments or from mitosis. Thus, a minor population of microtubules exists in cultured cells that contains an elevated level of tubulin modified in either one or two ways. While these two modifications occur primarily on the same subset of microtubules, they differ in their patterns of formation in vivo.

Acetylation↗

PubMed1997

Posttranslational modification of tubulin by palmitoylation: II. Identification of sites of palmitoylation.

As shown in the companion article, tubulin is posttranslationally modified in vivo by palmitoylation. Our goal in this study was to identify the palmitoylation sites by protein structure analysis. To obtain quantities of palmitoylated tubulin required for this analysis, a cell-free system for enzymatic [3H]palmitoylation was developed and characterized in our companion article. We then developed a methodology to examine directly the palmitoylation of all 451 amino acids of alpha-tubulin. 3H-labeled palmitoylated alpha-tubulin was cleaved with cyanogen bromide (CNBr). The CNBr digest was resolved according to peptide size by gel filtration on Sephadex LH60 in formic acid:ethanol. The position of 3H-labeled palmitoylated amino acids in peptides could not be identified by analysis of the Edman degradation sequencer product because the palmitoylated sequencer products were lost during the final derivatization step to phenylthiohydantoin derivatives. Modification of the gas/liquid-phase sequencer to deliver the intermediate anilinothiozolinone derivative, rather than the phenylthiohydantoin derivative, identified the cycle containing the 3H-labeled palmitoylated residue. Therefore, structure analysis of peptides obtained from gel filtration necessitated dual sequencer runs of radioactive peptides, one for sequence analysis and one to identify 3H-labeled palmitoylated amino acids. Further cleavage of the CNBr peptides by trypsin and Lys-C protease, followed by gel filtration on Sephadex LH60 and dual sequencer runs, positioned the 3H-labeled palmitoylated amino acid residues in peptides. Integration of all the available structural information led to the assignment of the palmitoyl moiety to specific residues in alpha-tubulin. The palmitoylated residues in alpha-tubulin were confined to cysteine residues only. The major site for palmitoylation was cysteine residue 376.

Amino Acid Sequence↗

PubMed1996

The apparent coupling between synthesis and posttranslational modification of Escherichia coli acyl carrier protein is due to inhibition of amino acid biosynthesis.

Acyl carrier protein (ACP) is modified on serine 36 by the covalent posttranslational attachment of 4'-phosphopantetheine from coenzyme A (CoA), and this modification is required for lipid biosynthesis. Jackowski and Rock (J. Biol. Chem 258:15186-15191, 1983) reported that upon depletion of the CoA pool by starvation for a CoA precursor, no accumulation of the unmodified form of ACP (apo-ACP) was detected. We report that this lack of apo-ACP accumulation results from decreased translation of the acpP mRNAs because of the limitation of the synthesis of glutamate and other amino acids made directly from tricarboxylic acid cycle intermediates.

Acyl Carrier Protein↗

PubMed1997

Effect of posttranslational modifications to myelin basic protein on its ability to aggregate acidic lipid vesicles.

When isolated from central nervous system myelin, myelin basic protein (MBP) exhibits charge microheterogeneity due to posttranslational deamidation, phosphorylation, and deimination of arginine to citrulline. These modifications are known to decrease the ability of MBP to aggregate acidic lipid vesicles and thus could regulate the ability of MBP to mediate adhesion between the intracellular surfaces of myelin. The effects of salt (KCl) concentration and the protein to lipid ratio on the ability of charge isomers of MBP to aggregate large unilamellar vesicles (LUVs) were investigated. Increased salt concentration from 10 to 100 mM caused increasing aggregation of LUVs by low concentrations of all charge isomers but did not eliminate the differences in their abilities to aggregate. All isomers were bound equally up to about 100 mM K+ but were dissociated at higher K+ concentrations. The degree of dissociation increased with increasing net negative charge of the isomer. At high concentrations all charge isomers except the form in which six arginine residues are converted to citrulline (C8) aggregated LUVs of phosphatidylcholine/phosphatidylserine (PC/PS) 8:2 (mol/mol) similarly and salt increased the aggregation to the same degree for all. There was less difference in the ability of the charge isomers, including C8, to aggregate LUVs with a lipid composition resembling that of the cytoplasmic leaflet of myelin (Cyt-LUVs) than for PC/PS LUVs. Furthermore, high salt concentrations (400 mM) did not dissociate any of the charge isomers from the Cyt-LUVs. These results suggest that the reason for inhibition of aggregating ability by charge modification is not increased charge repulsion of the protein but rather its reduced multivalency of net positive charge. They indicate further that the lipid composition of the cytoplasmic leaflet is ideally suited to permit MBP-mediated adhesion and that charge modifications of MBP would probably not affect adhesion of the intracellular surfaces of compact myelin where MBP concentration is high. However, charge modifications might affect adhesion in cytoplasm-containing regions of myelin such as the paranodal loops, where MBP concentration is low and where K+ concentration may vary in the range of 60-75 mM.

Animals↗

PubMed1993

Posttranslational modifications and assembly characteristics of goldfish tubulin.

Cell-free extracts from goldfish brain, ovarian follicles, testes and the cell line, ATCC CCL-71, were analyzed for posttranslationally modified tubulins. All samples, with the exception of that from brain where the reverse was true, contained more tyrosinated than detyrosinated alpha-tubulin. Additionally, extracts from brain and testes exhibited acetylated alpha-tubulin whereas this isoform was not visualized on blots of cell-free preparations from follicles and CCL-71 cells. Assembly of brain and ovary tubulin was induced with taxol. Brain tubulin, partially purified through three cycles of assembly/disassembly was associated with a variety of putative microtubule-associated proteins (MAPs), most of which had a high molecular mass. There were very few cold stable microtubules in brain preparations whereas, for ovary, purification of tubulin was hampered by significant losses of cold stable polymer. Comparison of brain and ovary showed there was no correlation between the extent of alpha-tubulin detyrosination or acetylation and cold stability of microtubules. Moreover, cycled tubulin from ovary contained acetylated tubulin even though this was not observed on blots of cell-free extracts from ovary or from follicles. Cultured goldfish cells contained extensive arrays of microtubules, many of which originated from discrete organizing centers. The results reveal the widespread distribution of posttranslationally modified tubulins in goldfish tissues, the differing assembly/disassembly characteristics of tubulin from brain versus ovary, and the presence of putative neural MAPs, mostly with a high molecular mass.

Animals↗

PubMed1989

Early posttranslational modifications of the three neurofilament subunits in mouse retinal ganglion cells: neuronal sites and time course in relation to subunit polymerization and axonal transport.

We have characterized stages in the posttranslational processing of the three neurofilament subunits, High (NF-H), Middle (NF-M), and Low (NF-L), in retinal ganglion cells in vivo during the interval between synthesis in cell bodies within the retina and appearance of these polypeptides in axons at the level of the optic nerve (optic axons). Neurofilament proteins pulse-labeled by injecting mice intravitreally with [35S]methionine or [32P]orthophosphate, were isolated from Triton-soluble and Triton-insoluble fractions of the retina or optic axons by immunoprecipitation or immunoaffinity chromatography. Within 2 h after [35S]methionine injection, the retina contained neurofilament-immunoreactive radiolabeled proteins with apparent molecular weights of 160, 139, and 70 kDa, which co-migrated with subunits of axonal neurofilaments that were dephosphorylated in vitro with alkaline phosphatase. The two larger polypeptides were not labeled with [32P]orthophosphate, indicating that they were relatively unmodified forms of NF-H and NF-M. About 75% of the subunits were Triton-insoluble by 2 h after isotope injection, and this percentage increased to 98% by 6 h. Labeled neurofilament polypeptides appeared in optic axons as early as 2 h after injection. These subunits exhibited apparent molecular weights of 160, 139, and 70 kDa and were Triton-insoluble. The time of appearance of fully modified polypeptide forms differed for each subunit (2 h for NF-L, 6-18 h for NF-M, 18-24 h for NF-H) and was preceded by the transient appearance of intermediate forms. The modified radiolabeled subunits in optic axons 3 days after synthesis were heavily labeled with [32P]orthophosphate and exhibited the same apparent molecular weights as subunits of axonal neurofilaments (70 kDa, 145 and 140 kDa, and 195-210 kDa, respectively). Whole mounts of retina immunostained with monoclonal antibodies against NF-H in different states of phosphorylation demonstrated a transition from non-phosphorylated neurofilaments to predominantly phosphorylated ones within a region of the axon between 200 and 1000 microns downstream from the cell body. These experiments demonstrate that the addition of most phosphate groups to NF-M and NF-H takes place within a proximal region of the axon. The rapid appearance of modified forms of NF-L after synthesis may imply that processing of this subunit occurs at least partly in the cell body. The presence of a substantial pool of Triton-insoluble, unmodified subunits early after synthesis indicates that the heaviest incorporation of phosphate occurs after neurofilament proteins are polymerized.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1995

Decreased molecular chaperone property of alpha-crystallins due to posttranslational modifications.

We studied the effect of oxidation, mixed disulfide formation and glycation of alpha-crystallins on their molecular chaperone property. The ability of alpha-crystallins to protect heat-induced denaturation and aggregation of beta L-crystallin was significantly diminished by these modifications. alpha-Crystallin from senile human lenses also showed significant loss of chaperone-like property. Age-dependent increase in posttranslationally modified alpha-crystallins is the likely cause for this change.

Adolescent↗

PubMed2005

Posttranslational modification of myxobacterial carrier protein domains in Pseudomonas sp. by an intrinsic phosphopantetheinyl transferase.

We demonstrate the ability of Pseudomonas putida KT2440, Pseudomonas syringae pv. tomato DC3000 and Pseudomonas stutzeri DSM10701 to posttranslationally activate carrier protein (CP) domains of various polyketide synthases, nonribosomal peptide synthetases, and fatty acid synthase by their intrinsic phosphopantetheinyl transferase. The apo-form is modified to the holo-form of the CP by attaching a phosphopantetheine moiety from coenzymeA to a conserved serine residue. The coding regions of the respective domains were cloned in order to generate C-terminal fusions with intein-chitin. The constructs were subcloned into a broad host range vector and transferred into the three pseudomonad hosts. The resulting recombinant pseudomonad strains were cultivated and each fusion protein was purified by affinity chromatography. Each purified CP was analysed using MALDI/TOF for the expected mass increase. Of the seven CPs tested, six could be purified from P. putida, which was chosen as the general host strain. Out of the six domains, five were completely activated, whereas only 5% of the protein of the sixth domain was in holo-form. Four domains were also expressed in the other hosts.

Amino Acid Sequence↗

PubMed1989

Posttranslational modifications of the beta subunit of a cryptomonad phycoerythrin. Sites of bilin attachment and asparagine methylation.

Determination of the partial amino acid sequence of the beta subunit of cryptomonad strain CBD phycoerythrin 566 established the nature, locations, and modes of attachment of the three bilin prosthetic groups and revealed a site of posttranslational methylation. Isolation of peptides cross-linked by a phycobiliviolin led to an unambiguous assignment of two thioether linkages, from residues beta-Cys-50 and beta-Cys-61 to this bilin. Two bilins were attached through single thioether linkages, a phycobiliviolin at beta-Cys-158 and a phycoerythrobilin at beta-Cys-82 (the residue numbering is that for B-phycoerythrin; Sidler, W., Kumpf, B., Suter, F., Morisset, W., Wehrmeyer, W., and Zuber, H. (1985) Biol. Chem. Hoppe-Seyler 366, 233-244). The partial sequences (99 residues) established for phycoerythrin 566 beta subunit showed a 79% identity with that of the red algal Porphyridium cruentum B-phycoerythrin beta subunit. A particularly remarkable finding is that the unique methylasparagine residue at position beta-72, highly conserved in cyanobacterial and red algal phycobiliproteins (Klotz, A. V., and Glazer, A. N. (1987) J. Biol. Chem. 262, 17350-17355), is also present at beta-72 in the cryptomonad phycoerythrin. Comparison of the locations of donor and acceptor bilins in cryptomonad phycoerythrin with those found for cyanobacterial and red algal phycobiliproteins showed different favored pathways of energy migration in the cryptomonad protein.

Amino Acid Sequence↗

PubMed1991

The herpes simplex virus 1 protein kinase encoded by the US3 gene mediates posttranslational modification of the phosphoprotein encoded by the UL34 gene.

Earlier studies have shown that a herpes simplex virus 1 (HSV-1) open reading frame, US3, encodes a novel protein kinase and have characterized the cognate amino acid sequence which is phosphorylated by this enzyme. This report identifies an apparently essential viral phosphoprotein whose posttranslational processing involves the viral protein kinase. Analyses of viral proteins phosphorylated in the course of productive infection revealed a phosphoprotein whose mobility was viral protein kinase and serotype dependent. Thus, the corresponding HSV-1 and HSV-2 phosphoproteins differ in their electrophoretic mobilities, and the phosphoprotein specified by the HSV-1 mutant deleted in US3 (R7041) differs from that of the corresponding HSV-1 and HSV-2 proteins. Analyses of HSV-1 x HSV-2 recombinants mapped the phosphoprotein between 0.42 and 0.47 map units on the prototype HSV-1 DNA map. Within this region, the UL34 open reading frame was predicted to encode a protein of appropriate molecular weight which would also contain the consensus target site for phosphorylation by the viral protein kinase as previously defined with synthetic peptides. Replacement of the native UL34 gene with a UL34 gene tagged with a 17-amino-acid epitope from the alpha 4 protein identified this gene as encoding the phosphoprotein. Finally, mutagenesis of the predicted phosphorylation site on UL34 in the viral genome, and specifically the substitution of threonine or serine with alanine in the product of the UL34 gene, yielded phosphoproteins whose electrophoretic mobilities could not be differentiated from that of the US3- mutant. We conclude that the posttranslational processing of the UL34 gene product to its wild-type phenotype requires the participation of the viral protein kinase. While the viral protein kinase is not essential for viral replication in cells in culture, the UL34 gene product itself may not be dispensable.

Amino Acid Sequence↗

PubMed2004

Posttranslational modification of brain tubulins from the Antarctic fish Notothenia coriiceps: reduced C-terminal glutamylation correlates with efficient microtubule assembly at low temperature.

We have shown previously that the tubulins of Antarctic fish assemble into microtubules efficiently at low temperatures (-2 to +2 degrees C) due to adaptations intrinsic to the tubulin subunits. To determine whether changes in posttranslational glutamylation of the fish tubulins may contribute to cold adaptation of microtubule assembly, we have characterized C-terminal peptides from alpha- and beta-tubulin chains from brains of adult specimens of the Antarctic rockcod Notothenia coriiceps by MALDI-TOF mass spectrometry and by Edman degradation amino acid sequencing. Of the four fish beta-tubulin isotypes, nonglutamylated isoforms were more abundant than glutamylated isoforms. In addition, maximal glutamyl side-chain length was shorter than that observed for mammalian brain beta tubulins. For the nine fish alpha-tubulin isotypes, nonglutamylated isoforms were also generally more abundant than glutamylated isoforms. When glutamylated, however, the maximal side-chain lengths of the fish alpha tubulins were generally longer than those of adult rat brain alpha chains. Thus, Antarctic fish adult brain tubulins are glutamylated differently than mammalian brain tubulins, resulting in a more heterogeneous population of alpha isoforms and a reduction in the number of beta isoforms. By contrast, neonatal rat brain tubulin possesses low levels of glutamylation that are similar to that of the adult fish brain tubulins. We suggest that unique residue substitutions in the primary structures of Antarctic fish tubulin isotypes and quantitative changes in isoform glutamylation act synergistically to adapt microtubule assembly to low temperatures.

Aging↗

PubMed1991

A combination of posttranslational modifications is responsible for the production of neuronal alpha-tubulin heterogeneity.

We describe the presence of alpha-tubulin and MAP2 acetyltransferase activities in mouse brain. The enzyme(s) copurified with microtubules through two cycles of assembly-disassembly. Incubation of microtubule proteins with [3H]acetyl CoA resulted in a strong labeling of both alpha-tubulin and MAP2. To determine the site of the modification, tubulin was purified and digested with Glu-C endoproteinase. A unique radioactive peptide was detected and purified by HPLC. Edman degradation sequencing showed that this peptide contained epsilon N-acetyllysine at position 40 of the alpha-tubulin molecule. This result demonstrates that mouse brain alpha-tubulin was acetylated at the same site as in Chlamydomonas. Isoelectric focusing analysis showed that acetylated alpha-tubulin was resolved into five isoelectric variants, denoted alpha 3 and alpha 5 to alpha 8. This heterogeneity is not due to acetylation of other sites but results from a single acetylation of Lys40 of an heterogeneous population of alpha-tubulin isoforms. These isoforms are produced by posttranslational addition of one to five glutamyl units. Thus, neuronal alpha-tubulin is extensively modified by a combination of modifications including acetylation, glutamylation, tyrosylation, and other yet unknown modifications.

Acetylation↗

PubMed1994

Posttranslational modification of an isoinhibitor from the potato proteinase inhibitor II gene family in transgenic tobacco yields a peptide with homology to potato chymotrypsin inhibitor I.

A member of the potato proteinase inhibitor II (PPI-II) gene family under the control of the cauliflower mosaic virus 35S promoter has been introduced into tobacco (Nicotiana tabacum). Purification of the PPI-II protein that accumulates in transgenic tobacco has confirmed that the N-terminal signal sequence is removed and that the inhibitor accumulates as a protein of the expected size (21 kD). However, a smaller peptide of approximately 5.4 kD has also been identified as a foreign gene product in transgenic tobacco plants. This peptide is recognized by an anti-PPI-II antibody, inhibits the serine proteinase chymotrypsin, and is not observed in nontransgenic tobacco. Furthermore, amino acid sequencing demonstrates that the peptide is identical to a lower molecular weight chymotrypsin inhibitor found in potato tubers and designated as potato chymotrypsin inhibitor I (PCI-I). Together, these data confirm that, as postulated to occur in potato, PCI-I does arise from the full-length PPI-II protein by posttranslational processing. The use of transgenic tobacco represents an ideal system with which to determine the precise mechanism by which this protein modification occurs.

Amino Acid Sequence↗

PubMed1999

A conotoxin from Conus textile with unusual posttranslational modifications reduces presynaptic Ca2+ influx.

Cone snails are gastropod mollusks of the genus Conus that live in tropical marine habitats. They are predators that paralyze their prey by injection of venom containing a plethora of small, conformationally constrained peptides (conotoxins). We report the identification, characterization, and structure of a gamma-carboxyglutamic acid-containing peptide, conotoxin epsilon-TxIX, isolated from the venom of the molluscivorous cone snail, Conus textile. The disulfide bonding pattern of the four cysteine residues, an unparalleled degree of posttranslational processing including bromination, hydroxylation, and glycosylation define a family of conotoxins that may target presynaptic Ca2+ channels or act on G protein-coupled presynaptic receptors via another mechanism. This conotoxin selectively reduces neurotransmitter release at an Aplysia cholinergic synapse by reducing the presynaptic influx of Ca2+ in a slow and reversible fashion. The three-dimensional structure, determined by two-dimensional 1H NMR spectroscopy, identifies an electronegative patch created by the side chains of two gamma-carboxyglutamic acid residues that extend outward from a cavernous cleft. The glycosylated threonine and hydroxylated proline enclose a localized hydrophobic region centered on the brominated tryptophan residue within the constrained intercysteine region.

1-Carboxyglutamic Acid↗

PubMed2006

Proteome analysis of human hair shaft: from protein identification to posttranslational modification.

The human hair proteome was investigated using two-dimensional LC-MS/MS. Among the 343 identified proteins, 70 were detected in high relative abundance, including keratin intermediate filament proteins, largely extractable with denaturants. Over 300 proteins were found to constitute the insoluble complex formed by transglutaminase cross-linking. The intracellular distribution of identified proteins is wide from cytoplasm to nucleus, mitochondria, ribosome, and plasma membrane. These results help rationalize ultrastructural features visible in the mature hair. Keratins and several substrates for transglutaminase were found to be posttranslationally modified by methylation and dimethylation. Evidence for ubiquitination of hair proteins was also obtained.

Amino Acid Sequence↗

PubMed1997

RNase Activity Decreases following a Heat Shock in Wheat Leaves and Correlates with Its Posttranslational Modification.

Heat shock results in a coordinate loss of translational efficiency and an increase in mRNA stability in plants. The thermally mediated increase in mRNA half-life could be a result of decreased expression and/or regulation of intracellular RNase enzyme activity. We have examined the fate of both acidic and neutral RNases in wheat seedlings that were subjected to a thermal stress. We observed that the activity of all detectable RNases decreased following a heat shock, which was a function of both the temperature and length of the heat shock. In contrast, no reduction in nuclease activity was observed following any heat-shock treatment. Antibodies raised against one of the major RNases was used in western analysis to demonstrate that the RNase protein level did not decrease following a heat shock, and the data suggest that the observed decrease in RNase activity in heat-shocked leaves may be due to modification of the protein. Two-dimensional gel/western analysis of this RNase revealed three isoforms. The most acidic isoform predominated in control leaves, whereas the most basic isoform predominated in leaves following a heat shock and correlated with the heat-shock-induced reduction in RNase activity and increase in mRNA half-life. These data suggest that RNase activity may be regulated posttranslationally following heat shock as a means to reduce RNA turnover until recovery ensues.

Journal Article↗

PubMed2005

Posttranslational modifications of p27kip1 determine its binding specificity to different cyclins and cyclin-dependent kinases in vivo.

Using 2-dimensional gel electrophoresis (2D-gel) analysis, we show here that cell-cycle entry is associated with a significant increase in p27(kip1) phosphorylation in human primary B cells. A similar pattern of increase in p27(kip1) phosphorylation was also seen in 2 fast-growing tumor cell lines, Burkitt lymphoma cell line BL40 and breast carcinoma cell line Cal51, where inactive p27(kip1) is expressed at high levels. Detailed analysis revealed for the first time that different cyclins and cyclin-dependent kinases (cdk's) interact with distinct posttranslationally modified isoforms of p27(kip1) in vivo. Cyclin E but not cyclin A selectively interacts with phosphorylated p27(kip1) isoforms, while cyclin D1 and D2 favor unphosphorylated p27(kip1) isoforms in vivo. Interestingly, cyclin D3 and cdk4 selectively interact with phosphorylated p27(kip1) in BL40 cells. Among all D-type cyclin/cdk4 and cdk6 complexes, cyclin D3/cdk4 is most active in sequestering the inhibitory activity of p27(kip1) in vitro in a cyclinE/cdk2 kinase assay. This novel feature of the binding specificity of p27(kip1) to cyclins and cdk's in vivo is interpreted in the context of overexpression of cyclin D3 in the presence of high levels of p27(kip1) in human B-cell lymphomas with adverse clinical outcome.

B-Lymphocytes↗