Peroxisomes are highly metabolic, autonomously replicating organelles that generate ROS as a by product of fatty acid β-oxidation. Consequently, cells must maintain peroxisome homeostasis, or risk pathologies associated with too few peroxisomes, such as peroxisome biogenesis disorders, or too many peroxisomes, inducing oxidative damage and promoting diseases such as cancer. We report that the PEX5 peroxisome import receptor binds ataxia-telangiectasia mutated (ATM) and localizes this kinase to the peroxisome. In response to reactive oxygen species (ROS), ATM signaling activates ULK1 and inhibits mTORC1 to induce autophagy. Specificity for autophagy of peroxisomes (pexophagy) is provided by ATM phosphorylation of PEX5 at Ser141, which promotes PEX5 mono-ubiquitination at K209, and recognition of ubiquitinated PEX5 by the autophagy adapter protein p62, directing the autophagosome to peroxisomes to induce pexophagy. These data reveal an important new role for ATM in metabolism as a sensor of ROS that regulates pexophagy.

SUMMARY Posttranslational modifications (PTMs) of tubulin specify microtubules for specialized cellular functions and comprise what is termed a “tubulin code”. PTMs of histones comprise an analogous “histone code”, although the “readers, writers and erasers” of the cytoskeleton and epigenome have heretofore been distinct. We show that methylation is a PTM of dynamic microtubules, and that the histone methytransferase, SETD2, which is responsible for H3 lysine 36 trimethylation (H3K36me3) of histones, also methylates α-tubulin at lysine 40, the same lysine that is marked by acetylation on microtubules. Methylation of microtubules occurs during mitosis and cytokinesis, and can be ablated by SETD2 deletion, which causes mitotic spindle and cytokinesis defects, micronuclei and polyploidy. These data now identify SETD2 as a dual function methyltransferase for both chromatin and the cytoskeleton, and show a requirement for methylation in maintenance of genomic stability and the integrity of both the tubulin and histone codes.

Subcellular localization is emerging as an important mechanism for mTORC1 regulation. We report that the tuberous sclerosis complex (TSC) signaling node, TSC1, TSC2 and Rheb, localizes to peroxisomes, where it regulates mTORC1 in response to reactive oxygen species (ROS). TSC1 and TSC2 were bound by PEX19 and PEX5, respectively, and peroxisome-localized TSC functioned as a Rheb GAP to suppress mTORC1 and induce autophagy. Naturally occurring pathogenic mutations in TSC2 decreased PEX5 binding, abrogated peroxisome localization, Rheb GAP activity, and suppression of mTORC1 by ROS. Cells lacking peroxisomes were deficient in mTORC1 repression by ROS and peroxisome-localization deficient TSC2 mutants caused polarity defects and formation of multiple axons in neurons. These data identify a role for TSC in Users may view, print, copy, download and text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms COMPETING FINANCIAL INTERESTSThe authors declare that they have no competing financial interests. HHS Public Access Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript responding to ROS at the peroxisome, and identify the peroxisome as a signaling organelle involved in regulation of mTORC1.Tuberous sclerosis complex (TSC) is a hereditary hamartoma syndrome caused by defects in either the TSC1 or TSC2 genes 1, 2 . The TSC tumor suppressor is a heterodimer comprised of tuberin (TSC2), a GTPase activating protein (GAP), and its activation partner hamartin (TSC1), which localizes the TSC tumor suppressor to endomembranes and protects TSC2 from proteasomal degradation 3,4 . TSC inhibits the activity of the small GTPase Rheb to repress mammalian target of rapamycin complex 1 (mTORC1) signaling, a negative regulator of autophagy [5][6][7][8][9][10][11][12] . mTORC1 is regulated by a variety of cellular stimuli including amino acids, mitogens such as insulin, glucose, and energy stress [13][14][15] . In the case of amino acids, which do not signal through TSC-Rheb pathway 15 , mTORC1 activity is regulated by the Rag GTPases, which form the Ragulator complex that localizes mTORC1 to the late endosome or lysosome compartment of cells [13][14][15][16][17][18] . We recently reported that TSC functions in a signaling node downstream of ataxia telangiectasia mutated (ATM) to repress mTORC1 in response to reactive oxygen species (ROS) 19 . However, identification of the specific subcellular compartment(s) in which the TSC tumor suppressor functions to regulate mTORC1 in response to ROS has heretofore remained elusive.Peroxisomes, carry out key metabolic functions in the cell including β-oxidation of fatty acids, and are a major source of cellular ROS 20,21 . Like mitochondria, peroxisomes are autonomously replicating organelles. Peroxisome biogenesis requires peroxin (PEX) proteins, which are essential for assembly of functional peroxisomes 22 . Specific PEX pro...

Two Sentrin/small ubiquitin-like modifier (SUMO)-specific protease 7 (SENP7) variants are naturally expressed in breast epithelia. Breast cancer (BCa) onset down-regulates the short SENP7 splice variant (SENP7S) and enhances the long transcript (SENP7L). Here, we show that SENP7L induction promotes gene expression profiles that favor aberrant proliferation and initiate epithelial-mesenchymal transition (EMT). SENP7L exhibits an interaction domain for the epigenetic remodeler heterochromatin protein 1 α (HP1α) and isopeptidase activity against SUMO-modified HP1α. Loss of this interaction domain, as observed with SENP7S, favors HP1α SUMOylation. SUMOylated HP1α is enriched at E2F-responsive and mesenchymal gene promoters, silences transcription of these genes, and promotes cellular senescence. Elevated SENP7L renders HP1α hypo-SUMOylated, which relieves transcriptional repression of the same genes and concurrently decreases transcription of epithelial-promoting genes via an HP1α-independent mechanism. Consequently, SENP7L levels correlate with EMT, motility, and invasiveness of BCa cells. Stable knockdown of elevated SENP7L levels lessens the dissemination of highly metastatic BCa cells to the lungs from primary implantation sites in in vivo studies. Thus, differential splicing of the SENP7 regulates either tumor suppression or progression.posttranslational-modification | epigenetics | dedifferentiation P osttranslational modification by the small ubiquitin-like modifier (SUMO) family has garnered much attention in cancer biology. This could be attributed to the ability of SUMOylation to elicit a rapid and reversible change to a protein's function and/or subcellular localization. However, SUMOylation of one cellular target may be critical for normal cell physiology, whereas SUMO conjugation of another protein may promote aberrant growth. Hence, targeting the SUMO-conjugating machinery may be detrimental to cancer as well as normal cells.Recently, the interest has shifted to the SUMO deconjugating enzymes or Sentrin/SUMO-specific proteases (SENP), which exhibit isopeptidase activity against a select subset of SUMOylated substrates. Although the expression of several SENPs is altered with onset of various carcinomas (1, 2), the physiological and/or pathophysiological function of many of the SENPs remains undefined. In addition, the six mammalian SENPs exhibit multiple splice variants that add to the functional diversity of the individual enzymes. SENP7 is an ideal example. Our laboratory used both extrinsic and experimental approaches to identify an SENP7 gene transcript with an intact catalytic domain (3). Recent studies show that the catalytic domain of SENP7 exhibits activity against synthetic and endogenous SUMO-conjugated substrates (4-6). Although these studies demonstrate SENP7's deSUMOylating capabilities, there exists a paucity of literature on the biological function of SENP7. In the present article we characterize two SENP7 transcript variants whose dueling functions support either tumor suppressive or po...

Tuberin, the Tsc2 gene product, integrates the phosphatidylinositol 3-kinase/mitogen-activated protein kinase (mitogenic) and LKB1/AMP-activated protein kinase (AMPK; energy) signaling pathways, and previous independent studies have shown that loss of tuberin is associated with elevated AMPK signaling and altered p27 function. In Tsc2-null tumors and tumor-derived cells from Eker rats, we observed elevated AMPK signaling and concordant cytoplasmic mislocalization of p27. Cytoplasmic localization of p27 in Tsc2-null cells was reversible pharmacologically using inhibitors of the LKB1/ AMPK pathway, and localization of p27 to the cytoplasm could be induced directly by activating AMPK physiologically (glucose deprivation) or genetically (constitutively active AMPK) in Tsc2-proficient cells. Furthermore, AMPK phosphorylated p27 in vitro on at least three sites including T170 near the nuclear localization signal, and T170 was shown to determine p27 localization in response to AMPK signaling. p27 functions in the nucleus to suppress cyclin-dependent kinase-2 (Cdk2) activity and has been reported to mediate an antiapoptotic function when localized to the cytoplasm. We found that cells with elevated AMPK signaling and cytoplasmic p27 localization exhibited elevated Cdk2 activity, which could be suppressed by inhibiting AMPK signaling. In addition, cells with elevated AMPK signaling and cytoplasmic p27 localization were resistant to apoptosis, which could be overcome by inhibition of AMPK signaling and relocalization of p27 to the nucleus. These data show that AMPK signaling determines the subcellular localization of p27, and identifies loss of integration of pathways controlling energy balance, the cell cycle, and apoptosis due to aberrant AMPK and p27 function as a feature of cells that have lost the Tsc2 tumor suppressor gene. [Cancer Res 2008;68(16):6496-506]