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Myristoylation, the N-terminal modification of proteins with the fatty acid myristate, is critical for membrane targeting and cell signaling. Because cancer cells often have increased N-myristoyltransferase (NMT) expression, NMTs were proposed as anti-cancer targets. To systematically investigate this, we performed robotic cancer cell line screens and discovered a marked sensitivity of hematological cancer cell lines, including B-cell lymphomas, to the potent pan-NMT inhibitor PCLX-001. PCLX-001 treatment impacts the global myristoylation of lymphoma cell proteins and inhibits early B-cell receptor (BCR) signaling events critical for survival. In addition to abrogating myristoylation of Src family kinases, PCLX-001 also promotes their degradation and, unexpectedly, that of numerous non-myristoylated BCR effectors including c-Myc, NFκB and P-ERK, leading to cancer cell death in vitro and in xenograft models. Because some treated lymphoma patients experience relapse and die, targeting B-cell lymphomas with a NMT inhibitor potentially provides an additional much needed treatment option for lymphoma.

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Myristoylation occurs cotranslationally on nascent proteins and post-translationally during apoptosis after caspase cleavages expose cryptic myristoylation sites. We demonstrate a drastic change in the myristoylated protein proteome in apoptotic cells, likely as more substrates are revealed by caspases. We show for the first time that both N-myristoyltransferases (NMTs) 1 and 2 are cleaved during apoptosis and that the caspase-3- or -8-mediated cleavage of NMT1 at Asp-72 precedes the cleavage of NMT2 by caspase-3 mainly at Asp-25. The cleavage of NMTs did not significantly affect their activity in apoptotic cells until the 8 h time point. However, the cleavage of the predominantly membrane bound NMT1 (64%) removed a polybasic domain stretch and led to a cytosolic relocalization (>55%), whereas predominantly cytosolic NMT2 (62%) relocalized to membranes when cleaved (>80%) after the removal of a negatively charged domain. The interplay between caspases and NMTs during apoptosis is of particular interest since caspases may not only control the rates of substrate production but also their myristoylation rate by regulating the location and perhaps the specificity of NMTs. Since apoptosis is often suppressed in cancer, the reduced caspase activity seen in cancer cells might also explain the higher NMT levels observed in many cancers.

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This article is available online at http://www.jlr.org "methyl-end" FA desaturases. Therefore, linoleic acid and ␣ -linolenic acid must be provided by the diet ( 3 ). These dietary essential FAs then serve as precursors for longer PUFAs of the n-6 and n-3 series, including arachidonic acid (20:4 n-6) and docosahexaenoic acid (22:6 n-3) required for many important physiological functions in humans ( 4,5 ). In this PUFA biosynthesis, only "front-end" desaturases ( ⌬ 5-and ⌬ 6-desaturases) are involved to introduce new double bonds between the preexisting double bonds and the carboxyl end of FAs ( 6 ).In 2000, the genomic structure of the fatty acid desaturase (FADS) cluster, located on chromosome 11 in humans, was reported ( 7 ). This cluster includes the FADS1 and FADS2 genes that code, respectively, for the well-known ⌬ 5-and ⌬ 6-desaturases involved in PUFA biosynthesis ( 8-10 ). A third gene was identifi ed on this cluster with high degree of nucleotide sequence homology with both FADS1 (62%) and FADS2 (70%) and was therefore named FADS3. Since its fi rst description in humans ( 7 ), FADS3 has been identifi ed in rats ( 11 ), baboons ( 12 ), mice ( 3 ), and many other mammals showing a similar nucleotide sequence and intron/exon organization ( 12 ). When analyzing the peptide sequence deduced from the FADS3 nucleotide sequence, FADS3 protein was identifi ed as a putative desaturase, according to its similarity with FADS1 and FADS2. Indeed, the predicted structure of FADS3 is composed of an N-terminal cytochrome b5-like domain characterized by a HPGG motif shown to be essential for the ⌬ 6-desaturase activity of FADS2 ( 13 ) QIEHH in the rat) characteristic of front-end-desaturases ( 14 ). According to its amino acid sequence, FADS3 was thereafter supposed to Abstract Fatty acid desaturases play critical roles in regulating the biosynthesis of unsaturated fatty acids in all biological kingdoms. As opposed to plants, mammals are so far characterized by the absence of desaturases introducing additional double bonds at the methyl-end site of fatty acids. However, the function of the mammalian fatty acid desaturase 3 (FADS3) gene remains unknown. This gene is located within the FADS cluster and presents a high nucleotide sequence homology with FADS1 ( ⌬ 5-desaturase) and FADS2 ( ⌬ 6-desaturase). Here, we show that rat FADS3 displays no common ⌬ 5-, ⌬ 6-or ⌬ 9-desaturase activity but is able to catalyze the unexpected ⌬ 13-desaturation of trans -vaccenate. Although there is no standard for complete conclusive identifi cation, structural characterization strongly suggests that the ⌬ 11,13-conjugated linoleic acid (CLA) produced by FADS3 from trans -vaccenate is the trans 11, cis 13-CLA isomer. In rat hepatocytes, knockdown of FADS3 expression specifically reduces trans -vaccenate ⌬ 13-desaturation. Evidence is presented that FADS3 is the fi rst "methyl-end" fatty acid desaturase functionally characterized in mammals. FA desaturases introduce double bonds between defi ned carbons of the fatty acyl chain and catalyze ...