Indiana University-Purdue University Indianapolis (IUPUI) / Glycogen is a major energy reserve in most eukaryotes and its rate of synthesis is controlled by glycogen synthase. The activity of eukaryotic glycogen synthase is regulated by the allosteric activator glucose-6-phosphate, which can overcome the inhibitory effects of phosphorylation. The effects of phosphorylation and glucose-6-phosphate on glycogen synthase are mediated by a cluster of six arginines located within a stretch of 12 amino acids near the C-terminus of the enzyme’s polypeptide chain. We studied isoform-2 of yeast glycogen synthase as a model to study the structural and molecular mechanisms that underlie the regulation of the eukaryotic enzymes and our primary tools of investigation were macromolecular X-ray crystallography, site-directed mutagenesis, intein-mediated peptide ligation and enzyme assays. We have solved the tetrameric structure of the yeast enzyme in two different activity states; the resting enzyme and the activated state when complexed with glucose-6-phosphate. Binding of glucose-6-phosphate to glycogen synthase induces large conformational changes that free the active site of the subunits to undergo conformational changes necessary to catalyze the reaction. Further, using site directed mutagenesis and intein-mediated peptide ligation to create specific phosphorylation states of the enzyme we were able to define specific roles for the arginine residues that mediate the regulatory effects of phosphorylation and glucose-6-phosphate activation. Based on these studies, we propose a three state structural model for the regulation of the enzyme, which relate the observed conformational states to specific activity levels. In addition to these regulatory studies, we have also solved the structure of the enzyme complexed with UDP and with substrate analogs, which provide detailed insight into the catalytic mechanism of the enzyme and the ability of glycogen synthase to remain tightly bound to its substrate glycogen.

The purpose of this study was to examine the effect of glucose ingestion on glycogen use andresynthesis during moderate exercise in untrained rats. Female Wistar rats (avg. wt.= 262 g) were assigned to either a control (C), control exercise (CE) or experimental exercise (EE) group. To examine glycogen resynthesis during exercise from a carbohydrate feeding, the EE animals were given 1 ml of a 10% glucose solution enriched with U-14C-glucose via stomach tube and run for 1h at 20.6 m/min. To distinguish between glycogen synthesis from endogenous versus exogenous sources, the CE animals were given a 0.1 ml tail-vein injection of U-14C-glucose tracer and run for 1h at 20.6 m/min. The C animals served as resting controls. Immediately after sacrifice, samples of the liver, soleus m., plantaris m., gastrocnemius m. and w. vastus m. were removed and analyzed for glycogen concentration and 14C activity in a glycogen pellet. Muscle and liver glycogen was 4.72 umol/g and 5.16 umol/g, respectively, higher (p<0.05) in the EE animals than in the CE animals. The average 14C activity of the muscle glycogen (mean ± SE) (182.85+ 31.79 cpm/g) was greater (p<0.05) than that of the liver (47.44 + 8.10 cpm/g), indicating a greater exogenous glucose incorporation into muscle glycogen than liver glycogen during exercise. However, this activity represented less than 2% of the remaining glycogen found in each tissue. Thus, it appears that the glycogen sparing observed with the glucose feeding in untrained rats was the result of an increased contribution of blood glucose to muscle metabolism.Ball State UniversityMuncie, IN 47306

Many of the physiological responses concomitant with exercise are understood. Similarly, many of the changes characterizing the aging process have been established. However, the combination of the two (ie. effects of aging on exercise or vice versa) presents a myriad of questions, of which many remain unanswered.The objective of this study was to establish the differences between previously untrained young and adult male Fischer 344 rats following an exhaustive swim for the following parameters: 1) muscle glycogen, an essential fuel substrate; 2) Glucose-6-phosphate dehydrogenase (G6PDH), a marker of inflammation and tissue damage; 3) citrate synthase (CS), an integral enzyme of the Kreb's cycle and a respiratory chain marker; 4) muscle protein; and 5) percent muscle dry weight.The rats were divided into two groups by age. Young (3 mo., n=16) and adult (12 mo., n=17) rats were randomly divided into sedentary (young sed (YSD) n=7 and adult sed (ASD) n=9) or exercised groups (young swimmers (YSW) n=8 and adult swimmers (ASW) n=8). Rats in the swimming groups were given a brief exposure to the water one week prior to their exhaustive swim to minimize the stress and confusion during the actual exercise bout. On the study days one randomly selected swimmer from each age group was swum to exhaustion and sacrificed via pneumothorax. One animal from each of the respective sedentary age groups was also randomly selected and sacrificed as above. The plantaris, rectus femoris, red vastus, soleus, triceps, and liver were surgically excised from each animal and frozen in liquid nitrogen for later analysis.While the younger animals had lower glycogen stores initially, following the exhaustive swim their reduction in muscle glycogen was approximately 150% that of the adult animals for any given muscle. Muscle glycogen levels in ASD and YSD rats were significantly higher than those of the YSW animals for all muscles with the exception of the YSD's soleus. However, the percent decrease in liver glycogen following the swim for the two age groups was almost identical (a reduction of 55.05% and 58.59% for the adult and young age groups, respectively).Although the adult animals were significantly heavier than the younger rats, this did not appear to cause a significant difference in their swim time to exhaustion. No significant differences were observed between the groups for muscle protein or G6PDH. Levels of CS were significantly higher in the YSD plantaris when compared to the ASW. Similarly, the ASD rectus femoris CS levels were significantly greater than those of the ASW. Although significant differences between groups in percent muscle dry weight existed for the plantaris, rectus femoris, and triceps such differences seemed to have little bearing on the two age group's swim to exhaustion times.On the basis of this study it was concluded that although starting with greater glycogen stores prior to exercise, adult animals use less of this substrate prior to exhaustion than do younger animals. While the mechanism for such a phenomenon was not discovered it is believed to be enzymatic in nature. Furthermore, the adult animals do not appear to exhibit significantly more tissue damage following an exhaustive swim than that seen in younger animals. / School of Physical Education

The purpose of this investigation was to determine the effect of passive and active recovery on the resynthesis of muscle glycogen after high intensity cycle ergometer exercise in untrained subjects. In a cross over design, six college-age males performed three, one min exercise bouts, at 130% V02max with a 4 min rest period between each work bout. Subjects refrained from exercise for two days prior to testing, and consumed a 15% carbohydrate solution (300g sugar in 2000ml of water) the day before each trial to help elevate glycogen concentration. The exercise protocol for each trial was identical, while the recovery following exercise was eitheractive (40-50% VO2max) or passive. The initial muscle glycogen values averaged 144.2 mmol•kg-1 w.w. for the active trial and 158.7 mmol•kg-1 w.w. for the passive trial. Corresponding post-exercise glycogen contents were 97.7 and 106.8 mmol•kg-1 w.w., respectively. These differences were not significant (P>0.05). However, the rate of muscle glycogen resynthesis during passive recovery increased 15 mmol•kg-1 w.w. whereas it decreased 6.27 mmol•kg-1 w.w. following active recovery (P<0.01). Also, the decrease in blood lactate concentration during active recovery was much faster than during passive recovery and significantly different at 10 and 30 min of the recovery period (P<0.01). The major finding of this investigation was that the rate of muscle glycogen resynthesis during passive recovery was significantly greater than that during active recovery. These data suggest that lactate can be used as an endogenous glycogenic precusor in muscle, and that glycogenesis was the prevalent pathway of lactate removal during passive recovery following high intensity cycle ergometer exercise. / Human Performance Laboratory

In an attempt to determine the extent to which well-trained endurance athletes practice the dietary recommendations for maximizing muscle glycogen resynthesis, twenty-four collegiate cross-country runners (14 males and 10 females) were chosen as subjects. The athletes kept four-day food and activity records during both a training and competitive period in the regular season. Energy intake was shown to be adequate in both phases. Total calories from carbohydrate, primarily complex, were found to be inadequate (<60%) for male runners and desirable (>60%) for females. Approximately 50% or less of the time carbohydrate was ingested immediately post-exercise, with even far less taken in suggested quantities (-1 g CHO/kg body weight). While the male athletes consumed primarily a combined solid and liquid form of carbohydrate immediately post-exercise, the females chose solid sources. Cereals and other breads were the most popular types of carbohydrate chosen immediately following exercise, in addition to commercial sports drinks/bars which were frequently ingested. An even more unfavorable trend in the distance runners was the infrequency of additional carbohydrate being ingested at two hour intervals following exercise. There were no significant differences in dietary trends between training and competitive phases. Overall these endurance athletes were not practicing the recommended feeding regimen for optimal muscle glycogen restoration. / Department of Home Economics

[Truncated abstract] It is well established that fish, amphibians and reptiles recovering from physical activity of near maximal intensity can replenish completely their muscle glycogen stores in the absence of food. In contrast, the extent to which these stores are replenished under these conditions in humans has been reported in all but one study to be partial. This implies that a few consecutive bouts of intense exercise might eventually lead to the sustained depletion of the muscle glycogen stores in humans if food is unavailable, thus limiting their capacity to engage in fight or flight behaviors unless mechanisms exist to protect muscle glycogen against sustained depletion. The objective of Study 1 was to test this prediction. Eight participants performed three intense exercise bouts each separated by a recovery period of 75 minutes. Although only 53% of muscle glycogen was replenished after the first exercise bout (postexercise and post-recovery glycogen levels of 246 ± 25 and 320 ± 36 mmol.kg-1 dry mass, respectively), all the glycogen mobilised during the second and third bouts was completely replenished during the respective recovery periods, with glycogen reaching levels of 319 ± 29 mmol.kg-1 dry mass after recovery from the third bout. These findings show that humans are not different from other vertebrate species in that there are conditions where humans have the ability to completely replenish without food intake the muscle glycogen mobilised during exercise. The results of our first study raise the intriguing possibility that humans have pre-set muscle glycogen levels that are protected against sustained depletion, with the extent to which muscle glycogen stores are replenished after exercise being dependent on the amount of glycogen required to attain those protected levels. ... During recovery, glycogen levels in the NORM group increased by more than ~50% and reached levels close to those alleged to be protected (189 ± 21 mmol.kg-1 dry mass), whereas no glycogen was deposited in the HCHO group. The sustained post-exercise activation of glycogen synthase, the transient fall in whole body carbohydrate oxidation rate, the increased mobilisation of body proteins, and the prolonged elevation in NEFA levels most probably played important roles in enabling glycogen synthesis in the NORM group. In conclusion, this thesis shows for the first time that there are some conditions (e.g. low pre-exercise muscle glycogen levels) where humans recovering from intense exercise have the capacity, like other species, to replenish completely their muscle glycogen stores from endogenous carbon sources. This study also suggests that humans protect preset levels of muscle glycogen against sustained depletion and at levels high enough to support at least one maximal sprint effort to exhaustion. Evidence is also provided for the existence of a feedback mechanism whereby glycogen below their protected levels mediate the activation of glycogen synthase to restore the depleted muscle glycogen stores back to their protected levels. Our findings, however, leave us with a number of novel unanswered questions which clearly show that the regulation of glycogen metabolism is far from the simple process generally depicted in most textbooks of biochemistry.

Thesis (MScMedSc)--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: INTRODUCTION: Glycogen synthase kinase-3 (GSK-3) is a serine-threonine protein kinase that was first discovered as a regulator of glycogen synthase thus playing a role in glycogen synthesis (Embi et al. 1980). GSK-3 has also been shown to down regulate the expression of SERCA-2a (a calcium ATPase pump) thus playing a role in myocardial contractility (Michael et al. 2004). However, SERCA-2a activity is regulated by phospholamban (PLM) and sarcolipin (SLN) (Asahi et al. 2003). GSK-3 is constitutively active in cells and can be acutely inactivated by insulin through phosphorylation by PKB/Akt. However, GSK-3 is known to phosphorylate and inhibit IRS-1 protein, thus disrupting insulin signaling (Eldar-Finkelman et al. 1996). In addition, abnormally high activities of GSK-3 protein has been implicated in several pathological disorders which include type 2 diabetes, neuron degenerative and affective disorders (Eldar-Finkelman et al 2009). This led to the development of new generations of inhibitors with specific clinical implications to treat these diseases (Martinez 2008). GSK-3 inhibition has been shown to improve insulin and blood glucose levels and to be cardioprotective during ischemia/reperfusion (Nikoulina et al. 2002; Kumar et al. 2007). AIMS: To determine whether myocardial GSK-3 protein and its substrate proteins are dysregulated in obesity and insulin resistance, and whether a specific GSK-3 inhibitor can prevent or reverse the cardiovascular pathology found in obese and insulin resistant animals. OBJECTIVES: To correlate the alterations in expression and activation of GSK-3 protein in a well characterised rat model of obesity coupled to insulin resistance with: i) myocardial contractile dysfunction and an inability of hearts to withstand ischemia/reperfusion, ii) the activation and expression of phospholamban and SERCA-2a in the sarcoplasmic reticulum, iii) the activation of intermediates (IRS-1, IRS-2 and PKB/Akt) that lie upstream in the activation pathway of GSK-3 and iv) to determine the effects of inhibition of GSK-3 on the abovementioned parameters. METHODS: Age and weight matched male Wistar rats (controls and diet induced obese (DIO) animals) were used in the present study. Controls were fed normal rat chow, while DIOs were fed a rat chow diet supplemented with sucrose and condensed milk, for 8 or 16 weeks. Half of each group of animals were treated with the GSK-3 inhibitor for 4 weeks (from 12 to 16 weeks). After the feeding and treatment period, animals were weighed, sacrificed, hearts removed and freeze clamped immediately or perfused with Krebs-Henseleit buffer and subjected to low flow ischemia (25 min) followed by 30 min reperfusion. Biometric (body weight, intraperitoneal fat, ventricular weight and tibia length) and biochemical (fasting blood glucose and insulin levels) parameters were determined. Expression of GSK-3, PKB/Akt, IRS-1, IRS-2, SERCA-2a and Phospholamban were determined by Western blotting. Ca2+ ATPase activity was determined spectrophotometrically. RESULTS: At both 8 and 16 weeks DIO animals were significantly bigger than control animals and this was associated with increased intraperitoneal fat in DIOs. In DIO animals: IRS-1 was downregulated at 8 weeks and both IRS-1 and IRS-2 as well as PKB/Akt at 16 weeks. There was an increased tendency of GSK-3 expression at both 8 and 16 weeks in DIO animals while SERCA-2a was severely downregulated from 8 weeks onwards and associated with lower Ca2+-ATPase activity. PLM expression was upregulated but its phosphorylation was attenuated. At 16 weeks, baseline heart rate (225 vs 275 in control, P<0.0001, n=6) and rate pressure product (21000 vs 30000 in control, P=0.019, n=6) were significantly lower in hearts from DIO animals. Functional recovery was unchanged but the time to ischemic contracture development was increased (11.6±0.4 control vs 16.2±0.5 min DIO, P<0.01, n=6). Treatment had no effect on total GSK-3 expression. However, GSK-3 phosphorylation was significantly increased in treated controls, while there was no significant difference in DIO animals. However, there was a tendency for an increased GSK-3 phosphorylation in treated DIO animals. GSK-3 inhibitor, improved hypertrophy in DIO animals, while it led to its development in control animals. GSK-3 inhibitor improved IRS-2 expression in both control and DIO animals while it had no effect on IRS-1 and SERCA-2a expression and activity. However, GSK-3 inhibition increased PKB/Akt and phospholamban phosphorylation in DIO animals. CONCLUSION: These findings show that high calorie diet as well as imbalance between energy intake and expenditure lead to the development of obesity and insulin resistance in male Wistar rats. We showed that GSK-3 and its substrate proteins are dysregulated in obesity and insulin resistance. The reduced SERCA-2a expression at baseline may have a negative impact on cardiac function. By treating the animals with GSK-3 inhibitor, we showed that GSK-3 protein may not be responsible for changes seen at baseline. The decreased IRS-1 and SERCA-2a expression may have been caused by a different mechanism other than the actions of GSK-3. However, according to this study, GSK-3 may play a role in regulation of IRS-2 expression but not in IRS-1. Increased PKB/Akt phosphorylation may contribute to the GSK-3 inhibition. In addition, GSK-3 inhibition may reverse cardiac hypertrophy in DIO animals, thus acting as a negative regulator of hypertrophy. / AFRIKAANSE OPSOMMING: Inleiding: Glikogeen sintase kinase-3 (GSK-3), 'n serien/threonien proteïen kinase, is oorspronklik ontdek as 'n rolspeler in glikogeen sintese, aangesien dit 'n reguleerder van glikogeen sintase is (Embi et al.1980). Intussen is dit ook bevind dat GSK-3 die uitdrukking van SERCA-2a ('n kalsium ATPase pomp) kan afreguleer en dus sodoende 'n rol speel in miokardiale kontraktiliteit (Michael et al. 2004). Die aktiwiteit van SERCA-2a kan egter ook gereguleer word deur fosfolamban (PLM) en sarkolipin (Asahi et al. 2003). GSK-3 is deurgaans aktief, maar kan tydelik geïnaktiveer word onder kondisies van insulien stimulasie deur PKB/Akt gemedieerde fosforilering. Aan die ander kant is dit bekend dat GSK-3 die IRS-1 proteïen kan fosforileer om dus sodoende insulien sein-transduksie af te reguleer (Eldar-Finkelman et al. 1996). Daarmee saam is abnormaal hoë vlakke van GSK-3 aktiwiteit geassosieer met verskeie patologiese versteurings, insluitend tipe 2 diabetes, neuron degeneratiewe en affektiewe versteurings (Eldar-Finkelman et al. 2009). Daar is dus nuwe generasies GSK-3 inhibitore ontwikkel met die kliniese potensiaal om hierdie patologieë te behandel (Martinez 2008). Dit is al bevind dat GSK-3 inhibisie geassosieer kan word met beide die normalisering van plasma insulien- en glukose vlakke, asook kardiobeskerming in die konteks van iskemie/herperfusie (Nikoulina et al. 2002; Kumar et al. 2007). Doelwitte: Om te bepaal of GSK-3 proteïen en sy substraat proteïene gedisreguleer is onder kondisies van obesiteit en insulien weerstandigheid, asook om vas te stel of 'n spesifieke GSK-3 inhibitor die kardiovaskulêre patologie wat gevind word in obese en insulien weerstandige diere kan verhoed of omkeer. Mikpunte: Om veranderinge in uitdrukking en aktiwiteit van GSK-3 proteïen in 'n goed gekarakteriseerde rotmodel van obesiteit, gekoppel aan insulien weerstandigheid, te korreleer met die volgende: i) miokardiale kontraktiele disfunksie en onvermoë om kardiale iskemie/herperfusie besering te weerstaan, ii) aktivering en uitdrukking van PLM en SERCA-2a in die sarkoplasmiese retikulum, iii) die aktivering van intermediêres wat proksimaal geleë is in die insulienseintransduksiepad van GSK-3 (IRS-1, IRS-2 en PKB/Akt) en iv) om die effek van behandeling met 'n spesifieke inhibitor van GSK-3 op die bogenoemde punte te bepaal. Metodes: Ouderdoms- en gewigsgepaarde manlike Wistar rotte (kontrole en dieet geïnduseerde obees (DIO) diere) is in die studie gebruik. Kontrole diere was normale rotkos gevoer, terwyl die DIO diere op 'n dieet van rotkos aangevul met sukrose en kondensmelk geplaas is vir 'n periode van 8 of 16 weke. Helfte van die diere van elke groep is behandel met die GSK-3 inhibitor vir 4 weke (vanaf week 12 tot 16). Na afloop van die voer- en behandelingsperiode is die diere geweeg, doodgemaak en die harte verwyder om dan of onmiddelik gevriesklamp te word, of retrograad geperfuseer te word met Krebs-Hensleit buffer. Ex vivo geperfuseerde harte is dan blootgestel aan 25 minute lae vloei iskemie gevolg deur 30 minute herperfusie. Biometriese (liggaamsgewig, intraperitoneale vet, ventrikulêre gewig en tibia lengte) en biochemiese (vastende bloedglukose en -insulien vlakke) parameters is telkens bepaal. Western klad tegnieke is gebruik om die uitdrukking en fosforilering van GSK-3, PKB/Akt, IRS-1, IRS-2, SERCA-2a en PLM te bepaal. Ca2+-ATPase aktiwiteit is spektrofotometries bepaal. Resultate: Na beide 8 en 16 weke was die DIO diere beduidend swaarder as die kontrole diere. Hierdie gewigstoename was geassosieer met meer intraperitoneale vet in die DIO diere. Verder, in die DIO diere was IRS-1 afgereguleer na 8 weke, terwyl beide IRS-1 en IRS-2 asook PKB/Akt afgereguleer was na 16 weke. GSK-3 uitdrukking het 'n neiging getoon om toe te neem na beide 8 en 16 weke in die DIO diere, terwyl SERCA-2a beduidend afgereguleer was reeds vanaf 8 weke, geassosieer met laer Ca2+-ATPase aktiwiteit. PLM uitdrukking het toegeneem en die fosforilering daarvan was verlaag. Op 16 weke was die basale harttempo (225 vs 275 in die kontrole groep, P<0.0001, n=6) en tempo druk produk (21000 vs 30000 in die kontrole groep, P=0.019, n=6) betekenisvol laer in die DIO diere. Funksionele herstel het onveranderd gebly, alhoewel die tyd tot iskemiese kontraktuur toegeneem het in die DIO groep (kontrole: 11.6±0.4 min vs DIO: 16.2±0.5 min, P<0.01, n=6). Toediening van die inhibitor het geen effek op totale GSK-3 uitdrukking gehad nie. Fosforilering van GSK-3 was egter wel beduidend verhoog in die behandelde kontrole diere, terwyl daar geen verskille in die DIO groep was nie. Die fosforilering van GSK-3 het wel geneig na 'n toename in die behandelde DIO diere. Die GSK-3 inhibitor het kontrasterende effekte op hipertrofie gehad: dit het dit omgekeer in die DIO groep, maar veroorsaak in die kontrole diere. Daarmee saam het die inhibitor die uitdrukking van IRS-2 in beide DIO en kontrole diere gestimuleer, maar geen effek op IRS-1 en SERCA-2a uitdrukking en aktiwiteit gehad nie. GSK-3 inhibisie het wel PKB/Akt en PLM fosforilering in die DIO diere verhoog. Gevolgtrekking: Hierdie bevindinge toon dat 'n hoë kalorie dieet, tesame met 'n wanbalans tussen energie inname en verbruiking, lei tot die ontwikkeling van obesiteit en insulien weerstand in manlike Wistar rotte. Die studie het ook getoon dat GSK-3 en sy substraat proteïene wel gedisreguleer is in obesiteit en insulien weerstandigheid. Die verlaagde basale uitdrukking van SERCA-2a mag dalk 'n negatiewe impak hê op kardiale funksie. Behandeling van die diere met 'n GSK-3 inhibitor het getoon dat GSK-3 moontlik nie verantwoordelik is vir die basislyn veranderinge nie. Die afname in IRS-1 en SERCA-2a uitdrukking kan moontlik toegeskryf word aan ander meganismes buiten die effekte van GSK-3. Hierdie studie toon wel dat GSK-3 moontlik 'n rol speel in die regulering van die uitdrukking van IRS-2, maar nie IRS-1 nie. Verhoogde PKB/Akt fosforilering mag dalk bydra tot die inhibisie van GSK-3. Daarmee saam blyk dit dat GSK-3 inhibisie hipertrofie kan omkeer in DIO diere, om dan sodoende op te tree as 'n negatiewe reguleerder van hipertrofie, maar in normale kontrole diere, hipertrofie in die hand werk. / South African Medical Research Council / University of Stellenbosch, Dept. of medical Physiology

Indiana University-Purdue University Indianapolis (IUPUI) / Sterol Regulatory Element Binding Protein-1 (SREBP-1) is a conserved transcription factor of the basic helix-loop-helix leucine zipper family (bHLH-Zip) that primarily regulates glycolytic and lipogenic enzymes such as L-pyruvate kinase, acetyl-CoA carboxylase, fatty acid synthase, stearoyl-CoA desaturase 1, and mitochondrial glycerol-3-phosphate acyltransferase 1. SREBP-1c activity is higher in the liver of human obese patients, as well as ob/ob and db/db mouse models of obesity and type 2 diabetes, underscoring the role of this transcription factor as a contributor to hepatic steatosis and insulin resistance. Nonetheless, SREBP-1 deficient ob/ob mice, do not display improved glycemia despite a significant decrease in hepatic lipid accumulation, suggesting that SREBP-1 might play a role at regulating carbohydrate metabolism. By silencing SREBP-1 in the liver of normal and type 2 diabetes db/db mice, we showed that indeed, SREBP-1 is needed for appropriate regulation of glycogen synthesis and gluconeogenesis enzyme gene expression. Depleting SREBP-1 activity more than 90%, resulted in a significant loss of glycogen deposition and increased expression of Pck1 and G6pc. Hence, the benefits of reducing de novo lipogenesis in db/db mice were offset by the negative impact on gluconeogenesis and glycogen synthesis. Some studies had also indicated that SREBP-1 regulates the insulin signaling pathway, through regulation of IRS2 and a subunit of the PI3K complex, p55g. To gain insight on the consequences of silencing SREBP-1 on insulin sensitivity, we analyzed the insulin signaling and mTOR pathways, as both are interconnected through feedback mechanisms. These studies suggest that SREBP-1 regulates S6K1, a downstream effector of mTORC1, and a key molecule to activate the synthesis of protein. Furthermore, these analyses revealed that depletion of SREBP-1 leads to reduced insulin sensitivity. Overall, our data indicates that SREBP-1 regulates pathways important for the fed state, including lipogenesis, glycogen and protein synthesis, while inhibiting gluconeogenesis. Therefore, SREBP-1 coordinates multiple aspects of the anabolic response in response to nutrient abundance. These results are in agreement with emerging studies showing that SREBP-1 regulates a complex network of genes to coordinate metabolic responses needed for cell survival and growth, including fatty acid metabolism; phagocytosis and membrane biosynthesis; insulin signaling; and cell proliferation.