Biomedical subjects

Alfred C O Vertegaal

Publications and source records attributed to Alfred C O Vertegaal.

4 recordsLinked to original sources

PubMed2006

Distinct and overlapping sets of SUMO-1 and SUMO-2 target proteins revealed by quantitative proteomics.

The small ubiquitin-like modifier (SUMO) family in vertebrates includes three different family members that are conjugated as post-translational modifications to target proteins. SUMO-2 and -3 are nearly identical but differ substantially from SUMO-1. We used quantitative proteomics to investigate the target protein preferences of SUMO-1 and SUMO-2. HeLa cells were established that stably express His6-SUMO-1 or His6-SUMO-2. These cell lines and control HeLa cells were labeled with stable arginine isotopes, and His6-SUMOs were enriched from lysates using immobilized metal affinity chromatography. 53 SUMO-conjugated proteins were identified, including 44 novel SUMO targets. 25 proteins were preferentially conjugated to SUMO-1, 19 were preferentially conjugated to SUMO-2, and nine proteins were conjugated to both SUMO-1 and SUMO-2. SART1 was confirmed by immunoblotting to have both SUMO-1- and SUMO-2-linked forms at similar levels. SUMO-1 and SUMO-2 are thus shown to have distinct and overlapping sets of target proteins, indicating that SUMO-1 and SUMO-2 may have both redundant and non-redundant cellular functions. Interestingly, 14 of the 25 SUMO-1-conjugated proteins contain zinc fingers. Although both SUMO family members play roles in many cellular processes, our data show that sumoylation is strongly associated with transcription because nearly one-third of the identified target proteins are putative transcriptional regulators.

Chromatography, Affinity↗

PubMed2004

A proteomic study of SUMO-2 target proteins.

The SUMO family in vertebrates includes at least three distinct proteins (SUMO-1, -2, and -3) that are added as post-translational modifications to target proteins. A considerable number of SUMO-1 target proteins have been identified, but little is known about SUMO-2. A stable HeLa cell line expressing His6-tagged SUMO-2 was established and used to label and purify novel endogenous SUMO-2 target proteins. Tagged forms of SUMO-2 were functional and localized predominantly in the nucleus. His6-tagged SUMO-2 conjugates were affinity-purified from nuclear fractions and identified by mass spectrometry. Eight novel potential SUMO-2 target proteins were identified by at least two peptides. Three of these proteins, SART1, heterogeneous nuclear ribonucleoprotein (RNP) M, and the U5 small nuclear RNP 200-kDa helicase, play a role in RNA metabolism. SART1 and heterogeneous nuclear RNP M were both shown to be genuine SUMO targets, confirming the validity of the approach.

Antigens, Neoplasm↗

PubMed2003

The N and C termini of the splice variants of the human mitogen-activated protein kinase-interacting kinase Mnk2 determine activity and localization.

The cap-binding eukaryotic initiation factor eIF4E is phosphorylated by the mitogen-activated protein (MAP) kinase-interacting kinases (Mnk's). Three forms of the Mnk's exist in human cells: Mnk1, Mnk2a, and Mnk2b. These last two are derived from the same gene by alternative splicing and differ only at their C termini. While Mnk2a contains a MAP kinase-binding site in this region, Mnk2b lacks such a sequence and is much less readily activated by MAP kinases in vitro. Expression of Mnk2b in mammalian cells leads to increased phosphorylation of eIF4E, showing that it acts as an eIF4E kinase in vivo. While Mnk2a is cytoplasmic, a substantial amount of Mnk2b is found in the nucleus. Both enzymes contain a stretch of basic residues in their N termini that plays a role in binding to eIF4G and functions as a nuclear localization signal. Binding of eIF4G or nuclear import appears to be regulated by the C terminus of Mnk2a. Furthermore, the MAP kinase-binding site of Mnk2a regulates nuclear entry. Within the nucleus, Mnk2b and certain variants of Mnk2a that are present in the nucleus colocalize with the promyelocytic leukemia protein PML, which also binds to eIF4E.

Alternative Splicing↗

PubMed2002

Differential expression of tapasin and immunoproteasome subunits in adenovirus type 5- versus type 12-transformed cells.

Adenovirus type 12 (Ad12)-transformed baby rat kidney (BRK) cells are oncogenic in syngeneic immunocompetent rats in contrast to adenovirus type 5 (Ad5)-transformed BRK cells, which are not oncogenic in these animals. A significant factor contributing to the difference in oncogenicity may be the low levels of major histocompatibility complex (MHC) class I membrane expression in Ad12-transformed BRK cells as compared with those in Ad5-transformed BRK cells, which presumably results in escape from killing by cytotoxic T lymphocytes. Here we show that, in addition to the decreased levels of expression of the MHC class I heavy chain and the peptide transporter Tap-2, the expression levels of the chaperone Tapasin and the immunoproteasome components MECL-1, PA28-alpha, and PA28-beta also are much lower in Ad12- than in Ad5-transformed BRK cells. The low expression levels of these proteins may contribute to the escape from killing by cytotoxic T lymphocytes, because the generation of optimal peptides and loading of these peptides on MHC class I require these components. Increased levels of phosphorylated signal transducer and activator of transcription-1 protein and expression of IFN regulatory factor-7 were found in Ad5- versus Ad12-transformed BRK cells. Therefore, the critical alteration leading to the plethora of differences may be an interferon (-related) effect.

ATP Binding Cassette Transporter, Subfamily B, Mem↗