Jaakko Mattila

FOXO-regulated transcription restricts overgrowth of Tsc mutant organs

FOXO is thought to function as a repressor of growth that is, in turn, inhibited by insulin signaling. However, inactivating mutations in Drosophila melanogaster FOXO result in viable flies of normal size, which raises a question over the involvement of FOXO in growth regulation. Previously, a growth-suppressive role for FOXO under conditions of increased target of rapamycin (TOR) pathway activity was described. Here, we further characterize this phenomenon. We show that tuberous sclerosis complex 1 mutations cause increased FOXO levels, resulting in elevated expression of FOXO-regulated genes, some of which are known to antagonize growth-promoting pathways. Analogous transcriptional changes are observed in mammalian cells, which implies that FOXO attenuates TOR-driven growth in diverse species.

Understanding Forkhead box class O function: lessons from Drosophila melanogaster.

Drosophila melanogaster is one of the most widely used model organisms. About 77% of known human disease genes have an ortholog in Drosophila, and many of the cellular signaling pathways are common between fruit flies and mammals. For example, a key signaling pathway in the regulation of growth and metabolism, the insulin/insulin-like growth factor 1 signaling pathway, is well conserved between flies and humans. Downstream effectors of this pathway are the Forkhead box class O (FOXO) family of transcription factors, with four members in mammals and a single FOXO protein in Drosophila, dFOXO. Research in Drosophila has been critical to elucidate the molecular mechanisms by which FOXO transcription factors regulate insulin signaling. In this review, we summarize the studies leading to dFOXO identification and its characterization as a central regulator of metabolism, life span, cell cycle, growth, and stress resistance.

RNAi screening for kinases and phosphatases identifies FoxO regulators

Forkhead box class O (FoxO) transcription factors are key regulators of growth, metabolism, life span, and stress resistance. FoxOs integrate signals from different pathways and guide the cellular response to varying energy and stress conditions. FoxOs are modulated by several signaling pathways, e.g., the insulin-TOR signaling pathway and the stress induced JNK signaling pathway. Here, we report a genome wide RNAi screen of kinases and phosphatases aiming to find regulators of dFoxO activity in Drosophila S2 cells. By using a combination of transcriptional activity and localization assays we identified several enzymes that modulate dFoxO transcriptional activity, intracellular localization and/or protein stability. Importantly, several currently known dFoxO regulators were found in the screening, confirming the validity of our approach. In addition, several interesting new regulators were identified, including protein kinase C and glycogen synthase kinase 3β, two proteins with important roles in insulin signaling. Furthermore, several mammalian orthologs of the proteins identified in Drosophila also regulate FOXO activity in mammalian cells. Our results contribute to a comprehensive understanding of FoxO regulatory processes.

Drosophila FoxO Regulates Organism Size and Stress Resistance through an Adenylate Cyclase

ABSTRACT Forkhead box class O (FoxO) transcription factors are a family of conserved proteins that regulate the cellular responses to various stimuli, such as energy deprivation, stress, and developmental cues. FoxO proteins are important mediators of the insulin signaling pathway, adjusting growth and metabolism to nutrient availability. Insulin signaling acts together with the glucagon-stimulated cAMP signaling pathway to orchestrate the organism response to various nutritional conditions. In this study, we demonstrate that Drosophila melanogaster FoxO (dFoxO) regulates cAMP signaling by directly inducing the expression of an adenylate cyclase gene, ac76e. Interestingly, ac76e is expressed in a highly restricted pattern throughout fly development, limited to the corpus allatum (CA), gastric cecum, and malpighian tubules. dFoxO activation of AC76E in the CA increases starvation resistance and limits growth. Our results unravel a new role for dFoxO, integrating cAMP and insulin signaling to adapt organism growth to the existing nutritional conditions.

GMF Promotes Leading-Edge Dynamics and Collective Cell Migration In Vivo

Lamellipodia are dynamic actin-rich cellular extensions that drive advancement of the leading edge during cell migration. Lamellipodia undergo periodic extension and retraction cycles, but the molecular mechanisms underlying these dynamics and their role in cell migration have remained obscure. We show that glia-maturation factor (GMF), which is an Arp2/3 complex inhibitor and actin filament debranching factor, regulates lamellipodial protrusion dynamics in living cells. In cultured S2R(+) cells, GMF silencing resulted in an increase in the width of lamellipodial actin filament arrays. Importantly, live-cell imaging of mutant Drosophila egg chambers revealed that the dynamics of actin-rich protrusions in migrating border cells is diminished in the absence of GMF. Consequently, velocity of border cell clusters undergoing guided migration was reduced in GMF mutant flies. Furthermore, genetic studies demonstrated that GMF cooperates with the Drosophila homolog of Aip1 (flare) in promoting disassembly of Arp2/3-nucleated actin filament networks and driving border cell migration. These data suggest that GMF functions in vivo to promote the disassembly of Arp2/3-nucleated actin filament arrays, making an important contribution to cell migration within a 3D tissue environment.

Mondo-Mlx Mediates Organismal Sugar Sensing through the Gli-Similar Transcription Factor Sugarbabe

The ChREBP/Mondo-Mlx transcription factors are activated by sugars and are essential for sugar tolerance. They promote the conversion of sugars to lipids, but beyond this, their physiological roles are insufficiently understood. Here, we demonstrate that in an organism-wide setting in Drosophila, Mondo-Mlx controls the majority of sugar-regulated genes involved in nutrient digestion and transport as well as carbohydrate, amino acid, and lipid metabolism. Furthermore, human orthologs of the Mondo-Mlx targets display enrichment among gene variants associated with high circulating triglycerides. In addition to direct regulation of metabolic genes, Mondo-Mlx maintains metabolic homeostasis through downstream effectors, including the Activin ligand Dawdle and the Gli-similar transcription factor Sugarbabe. Sugarbabe controls a subset of Mondo-Mlx-dependent processes, including de novo lipogenesis and fatty acid desaturation. In sum, Mondo-Mlx is a master regulator of other sugar-responsive pathways essential for adaptation to a high-sugar diet.

Bilateral Ectasia After Femtosecond Laser-Assisted Small Incision Lenticule Extraction (SMILE)

PURPOSE To describe a case of bilateral ectasia after small incision lenticule extraction (SMILE) in a patient with early keratoconus. METHODS Case report. RESULTS Bilateral SMILE was performed on a patient even though preoperative topographies showed changes indicating early keratoconus. The right eye underwent further photorefractive keratectomy enhancement 18 months later. The patient developed a bilateral corneal ectasia. CONCLUSIONS This case underlines the importance of thorough preoperative assessment for possible keratoconus suspect changes with corneal topography to avoid postoperative ectasia. [J Refract Surg. 2016;32(7):497-500.].

Treatment of Pseudomonas aeruginosa keratitis with combined corneal cross-linking and human amniotic membrane transplantation

0.01) compared with vehicle treatment (14.13 ± 1.73%, n = 14, Fig. 1A, B). By contrast, the ratio of normal vascularized area is higher in the animals treated with VAP-1 inhibitor (94.69 ± 0.82%, n = 18, p < 0.01) than those treated with vehicle (83.56 ± 1.56%, n = 14, Fig. 1C), indicating that VAP-1 blockade did not affect physiological vascular development in OIR model. The present data demonstrated that VAP-1 blockade selectively prevents pathological retinal neovascularization without inhibiting physiological neovascularization. Previous data demonstrated that VAP-1 blockade attenuated the angiogenic response via suppression of recruitment of monocyte lineage cells (Noda et al. 2008; Nakao et al. 2011). As it has been elucidated that monocyte lineage cells are involved in the pathological neovascularization in OIR model (Ishida et al. 2003), it is likely that VAP-1 blockade attenuates the pathological neovascularization through suppression of macrophage recruitment. Interestingly, the current data also demonstrated that physiological neovascularization was not inhibited and rather increased in VAP-1 inhibitortreated group compared with vehicletreated group. It was reported that VAP-1 is expressed from embryonic day 14 in mouse retina, which may indicate that VAP-1 plays a role for vascularization during eye development (Valente et al. 2008). However, the current data showed that VAP-1 blockade did not suppress physiological neovascularization in OIR model and therefore suggested that the role of VAP-1 might not be identical in physiological and pathological angiogenesis. In the treatment of ROP, the therapeutic agents that suppress pathological, but not physiological, neovascularization are ideal. Further studies to evaluate the effect of VAP-1 blockade in OIR model are warranted.

Penetrating Keratoplasty for Corneal Amyloidosis in Familial Amyloidosis, Finnish Type

PURPOSE To analyze the outcome of penetrating keratoplasty (PK) to the first eye for corneal amyloidosis in familial amyloidosis, Finnish type (FAF). DESIGN Single-center, retrospective, nonrandomized, interventional, noncomparative case series. PARTICIPANTS Thirty-one eyes of 31 patients with FAF. INTERVENTION All patients with FAF who had their first PK in Helsinki University Eye Hospital between January 1, 1990, and August 1, 2011, were identified and a retrospective analysis of the patient charts was performed. MAIN OUTCOME MEASURES Best spectacle-corrected visual acuity (BCVA), intraoperative and postoperative complications, graft survival, reason for graft failure, and frequency of regrafting. RESULTS The median follow-up period was 32 months (range, 5-114). After 24 months, the median BCVA was 1.15 on a logarithm of the minimum angle of resolution scale (20/280; mean, 1.1; SD, 0.5) in comparison with the preoperative median BCVA of 1.3 (20/400; mean, 1.3; SD, 0.4). At 24 months, 3 of 18 eyes (17%) had a visual acuity of ≥0.5 (20/63) and 13 of 18 grafts (72%) were clear. Rejection occurred in 6 of 31 primary grafts (19%). Graft failure occurred in 16 of 31 eyes and resulted from surface complications in 11 eyes and additionally from rejection in 5 eyes. Seven eyes needed regrafting (twice in 1 eye). Complications were frequent in the early and late postoperative periods. Presence of preoperative corneal or graft neovascularization was an indicator of a high risk of graft failure and poor visual outcome. CONCLUSIONS In a minority of FAF patients, PK improves vision. Owing to the high failure risk and guarded visual prognosis after PK, it is important that both the surgeon and the patient have realistic expectations. It may be reasonable to limit PK to cases with bilateral advanced disease. It seems reasonable to optimize ocular surface health and to delay PK.

Insights to transcriptional networks by using high throughput RNAi strategies

RNA interference (RNAi) is a powerful method to unravel the role of a given gene in eukaryotic cells. The development of high throughput assay platforms such as fluorescence plate readers and high throughput microscopy has allowed the design of genome wide RNAi screens to systemically discern members of regulatory networks around various cellular processes. Here we summarize the different strategies employed in RNAi screens to reveal regulators of transcriptional networks. We focus our discussion in experimental approaches designed to uncover regulatory interactions modulating transcription factor activity.