Insights into the molecular mechanisms and signalling pathways of epithelial to mesenchymal transition (EMT) in colorectal cancer: A systematic review and bioinformatic analysis of gene expression.

Colorectal cancer (CRC) is ranked as the second leading cause of mortality worldwide, mainly due to metastasis. Epithelial to mesenchymal transition (EMT) is a complex cellular process that drives CRC metastasis, regulated by changes in EMT-associated gene expression. However, while numerous genes have been identified as EMT regulators through various in vivo and in vitro studies, little is known about the genes that are differentially expressed in CRC tumour tissue and their signalling pathway in regulating EMT. Using an integration of systematic search and bioinformatic analysis, gene expression profiles of CRC tumour tissues were compared to non-tumour adjacent tissues to identify differentially expressed genes (DEGs), followed by performing systematic review on common identified DEGs. Fifty-eight common DEGs were identified from the analysis of 82 tumour tissue samples obtained from four gene expression datasets (NCBI GEO). These DEGS were then systematically searched for their roles in modulating EMT in CRC based on previously published studies. Following this, 10 common DEGs (CXCL1, CXCL8, MMP1, MMP3, MMP7, TACSTD2, VIP, HPGD, ABCG2, CLCA4) were included in this study and subsequently subjected to further bioinformatic analysis. Their roles and functions in modulating EMT in CRC were discussed in this review. This study enhances our understanding of the molecular mechanisms underlying EMT and uncovers potential candidate genes and pathways that could be targeted in CRC.

Interleukin-1 alpha and high mobility group box-1 secretion in polyinosinic:polycytidylic-induced colorectal cancer cells occur via RIPK1-dependent mechanism and participate in tumourigenesis.

Pathogenic infections have significant roles in the pathogenesis of colorectal cancer (CRC). These infections induce the secretion of various damage-associated molecular patterns (DAMPs) including interleukin-1 alpha (IL-1α) and high mobility group box-1 (HMGB1). Despite their implication in CRC pathogenesis, the mechanism(s) that modulate the secretion of IL-1α and HMGB1, along with their roles in promoting CRC tumourigenesis remain poorly understood. To understand the secretory mechanism, HT-29 and SW480 cells were stimulated with infectious mimetics; polyinosinic:polycytidylic acid [Poly(I:C)], lipopolysaccharide (LPS) and pro-inflammatory stimuli; tumour necrosis factor-alpha (TNF-α). IL-1α and HMGB1 secretion levels upon stimulation were determined via ELISA. Mechanism(s) mediating IL-1α and HMGB1 secretion in CRC cells were characterized using pharmacological inhibitors and CRISPR-Cas9 gene editing targeting relevant pathways. Recombinant IL-1α and HMGB1 were utilized to determine their impact in modulating pro-tumourigenic properties of CRC cells. Pharmacological inhibition showed that Poly(I:C)-induced IL-1α secretion was mediated through endoplasmic reticulum (ER) stress and RIPK1 signalling pathway. The secretion of HMGB1 was RIPK1-dependent but independent of ER stress. RIPK1-targeted CRC cell pools exhibited decreased cell viability upon Poly(I:C) stimulation, suggesting a potential role of RIPK1 in CRC cells survival. IL-1α has both growth-promoting capabilities and stimulates the production of pro-metastatic mediators, while HMGB1 only exhibits the latter; with its redox status having influence. We demonstrated a potential role of RIPK1-dependent signalling pathway in mediating the secretion of IL-1α and HMGB1 in CRC cells, which in turn enhances CRC tumorigenesis. RIPK1, IL-1α and HMGB1 may serve as potential therapeutic targets to mitigate CRC progression.

Metabolic changes in vitamin D receptor knockout mice.

VDR expression has been found in many cell types involved in metabolism, including the beta-cells of the pancreatic islets. Activated vitamin D and its interactions with the vitamin D receptor (VDR) are implicated in glucose homeostasis. We investigated the metabolic phenotype of the VDR-null (VDRKO) mouse at early and middle age. All offspring of heterozygote VDRKO breeding-pairs were fed 'rescue diet' from weaning to normalize calcium and phosphate levels in VDRKO and to avoid confounding by different diets. Glucose tolerance testing was performed at 7 and 24 weeks of age. Insulin tolerance testing, glucose-stimulated insulin secretion, body-composition studies and islet isolation were performed at 25-27 weeks. Glucose-stimulated insulin secretion was tested in isolated islets. VDRKO mice had reduced bone density, subcutaneous fat mass and muscle weights compared to WT mice. Despite reduced fat mass, glucose tolerance did not differ significantly. Male but not female VDRKO had improved insulin sensitivity. Global loss of VDR has significant effects on organs involved in energy metabolism and glucose homeostasis. In the setting of decreased fat mass, a clear effect on glucose tolerance was not present.

Iron chelation increases beige fat differentiation and metabolic activity, preventing and treating obesity.

Beige and brown fat consume glucose and lipids to produce heat, using uncoupling protein 1 (UCP1). It is thought that full activation of brown adipose tissue (BAT) may increase total daily energy expenditure by 20%. Humans normally have more beige and potentially beige-able fat than brown fat. Strategies to increase beige fat differentiation and activation may be useful for the treatment of obesity and diabetes. Mice were fed chow or high-fat diet (HFD) with or without the iron chelator deferasirox. Animals fed HFD + deferasirox were markedly lighter than their HFD controls with increased energy expenditure (12% increase over 24 h, p < 0.001). Inguinal fat from HFD + deferasirox mice showed increased beige fat quantity with greater Ucp1 and Prdm16 expression. Inguinal adipose tissue explants were studied in a Seahorse bioanalyser and energy expenditure was significantly increased. Deferasirox was also effective in established obesity and in ob/ob mice, indicating that intact leptin signalling is not needed for efficacy. These studies identify iron chelation as a strategy to preferentially activate beige fat. Whether activating brown/beige fat is effective in humans is unproven. However, depleting iron to low-normal levels is a potential therapeutic strategy to improve obesity and related metabolic disorders, and human studies may be warranted.

Emergence of antibiotic-resistant Mycoplasma genitalium as the cause of non-gonococcal urethritis in male patients at a sexually transmitted infection clinic.

Mycoplasma genitalium is the cause of an emerging sexually transmitted infection (STI) with high propensity for development of antimicrobial resistance. In a prevalence study conducted at the public STI service in Hong Kong, the first void urine samples of 38 (8%) of 493 male patients with non-gonococcal urethritis (NGU) tested positive for M. genitalium using reverse transcription polymerase chain reaction. Patients with M. genitalium infection were younger [31 vs 33 years, odds ratio (OR) 0.96, 95% confidence interval (CI) 0.93-0.996; P=0.03], more likely to present with urethral discharge (12% vs 6%, OR 2.16, 95% CI 1.10-4.23; P=0.02) and had symptom duration >2 weeks (14% vs 6%, OR 2.34, 95% CI 1.10-4.97; P=0.03) compared with patients without M. genitalium infection. The prevalence of M. genitalium infection was lower in patients co-infected with Chlamydia trachomatis compared with patients with isolated infection (4% vs 10%, OR 0.38, 95% CI 0.17-0.84; P=0.02). The prevalence of M. genitalium infection was not higher in men who have sex with men. Antimicrobial-resistance-conferring mutations were present in 24 (63%) patients with M. genitalium - 23S rRNA 18 (47%) and parC 19 (53%). Similar to neighbouring countries in the Asia Pacific region, concurrent resistance mutations against both macrolides and fluoroquinolones were demonstrated in 14 (37%) patients. Histories of azithromycin and moxifloxacin use were significantly associated with a diagnosis of M. genitalium infection. Characteristically, NGU in Hong Kong featured the co-existence of mono-resistance against macrolides or fluoroquinolones, and the presence of dual class resistance. The geographic variability of antimicrobial resistance against M. genitalium is attributed not just to the different transmission networks formed in separate population groups, but the antimicrobial prescriptions for the treatment of urethritis in the community.

IL-1α and colorectal cancer pathogenesis: Enthralling candidate for anti-cancer therapy.

Inflammation has been well-established as a hallmark of colorectal cancer (CRC). Interleukin-1 alpha (IL-1α) is one of the primary inflammatory mediators driving the pathogenesis of inflammation-associated CRC. This systematic review presents the roles of IL-1α in the pathogenesis of the disease. Bibliographic databases PubMed, Science Direct, Scopus and Web of Science were systematically searched for articles that addresses the relationship between IL-1α and colorectal cancer. We highlighted various mechanisms by which IL-1α promotes the pathogenesis of CRC including enhancement of angiogenesis, metastasis, resistance to therapy, and inhibition of tumour suppressive genes. We also discussed the potential mechanisms by which IL-1α expression is induced or secreted in various studies. Beyond these, the systematic review also highlights several potential therapeutic strategies which should be further explored in the future; to target IL-1α and/or its associated pathways; paving our way in finding effective treatments for CRC patients.

HMGB1: an overview of its versatile roles in the pathogenesis of colorectal cancer.

BACKGROUND: In recent years, the high mobility group box-1 (HMGB1) protein, a damage-associated molecular pattern (DAMP) molecule, has been found to play multifunctional roles in the pathogenesis of colorectal cancer. Although much attention has been given to the diagnostic and prognostic values of HMGB1 in colorectal cancer, the exact functional roles of the protein as well as the mechanistic pathways involved have remained poorly defined. This systematic review aims to discuss what is currently known about the roles of HMGB1 in colorectal cancer development, growth and progression, and to highlight critical areas for future investigations. To achieve this, the bibliographic databases Pubmed, Scopus, Web of Science and ScienceDirect were systematically screened for articles from inception till June 2018, which address associations of HMGB1 with colorectal cancer. CONCLUSIONS: HMGB1 plays multiple roles in promoting the pathogenesis of colorectal cancer, despite a few contradicting studies. HMGB1 may differentially regulate disease-related processes, depending on the redox status of the protein in colorectal cancer. Binding of HMGB1 to various protein partners may alter the impact of HMGB1 on disease progression. As HMGB1 is heavily implicated in the pathogenesis of colorectal cancer, it is crucial to further improve our understanding of the functional roles of HMGB1 not only in colorectal cancer, but ultimately in all types of cancers.

Hepatic Aryl hydrocarbon Receptor Nuclear Translocator (ARNT) regulates metabolism in mice.

BACKGROUND & AIMS: Aryl hydrocarbon Receptor Nuclear Translocator (ARNT) and its partners hypoxia-inducible factors (HIF)-1α and HIF-2α are candidate factors for the well-known link between the liver, metabolic dysfunction and elevation in circulating lipids and glucose. Methods: Hepatocyte-specific ARNT-null (LARNT), HIF-1α-null (LHIF1α) and HIF-2α-null (LHIF2α) mice were created. RESULTS: LARNT mice had increased fasting glucose, impaired glucose tolerance, increased glucose production, raised post-prandial serum triglycerides (TG) and markedly lower hepatic ATP versus littermate controls. There was increased expression of G6Pase, Chrebp, Fas and Scd-1 mRNAs in LARNT animals. Surprisingly, LHIF1α and LHIF2α mice exhibited no alterations in any metabolic parameter assessed. CONCLUSIONS: These results provide convincing evidence that reduced hepatic ARNT can contribute to inappropriate hepatic glucose production and post-prandial dyslipidaemia. Hepatic ARNT may be a novel therapeutic target for improving post-prandial hypertriglyceridemia and glucose homeostasis.

Transplantation sites for human and murine islets.

AIMS/HYPOTHESIS: Beta cell replacement is a potential cure for type 1 diabetes. In humans, islet transplants are currently infused into the liver via the portal vein, although this site has disadvantages. Here, we investigated alternative transplantation sites for human and murine islets in recipient mice, comparing the portal vein with quadriceps muscle and kidney, liver and spleen capsules. METHODS: Murine islets were isolated from C57BL6/J mice and transplanted into syngeneic recipients. Human islets were isolated and transplanted into either severe combined immunodeficiency (SCID) or recombination-activating gene 1 (RAG-1) immunodeficient recipient mice. All recipient mice were 8-12 weeks of age and had been rendered diabetic (defined as blood glucose concentrations ≥20 mmol/l on two consecutive days before transplantation) by alloxan tetrahydrate treatment. Islets were transplanted into five different sites (portal vein, quadriceps muscle, kidney, liver and spleen capsules). Blood glucose concentrations were monitored twice weekly until mice were killed. Dose-response studies were also performed to determine the minimum number of islets required to cure diabetes ('cure' is defined for this study as random fed blood glucose of <15 mmol/l). RESULTS: For transplantation of murine islets into the different sites, the kidney yielded 100% success, followed by muscle (70%), portal vein (60%), spleen capsule (29%) and liver capsule (0%). For human islets, transplantation into the kidney cured diabetes in 75-80% of recipient mice. Transplantation into muscle and portal vein had intermediate success (both 29% at 2000 islet equivalents), while transplantation into liver and spleen capsule failed (0%). With increased islet mass, success rates for muscle grafts improved to 52-56%. CONCLUSIONS/INTERPRETATION: For both human and murine islets, equivalent or superior glucose lowering results were obtained for transplantation into skeletal muscle, compared with the portal vein. Unfortunately, kidney grafts are not feasible in human recipients. Skeletal muscle offers easier access and greater potential for protocol biopsies. This study suggests that human trials of muscle as a transplant site may be warranted.

Vitamin D signaling regulates proliferation, differentiation, and myotube size in C2C12 skeletal muscle cells.

Vitamin D deficiency is linked to a range of muscle disorders including myalgia, muscle weakness, and falls. Humans with severe vitamin D deficiency and mice with transgenic vitamin D receptor (VDR) ablation have muscle fiber atrophy. However, molecular mechanisms by which vitamin D influences muscle function and fiber size remain unclear. A central question is whether VDR is expressed in skeletal muscle and is able to regulate transcription at this site. To address this, we examined key molecular and morphologic changes in C2C12 cells treated with 25-hydroxyvitamin D (25OHD) and 1,25-dihydroxyvitamin D (1,25(OH)(2)D). As well as stimulating VDR expression, 25(OH)D and 1,25(OH)(2)D dose-dependently increased expression of the classic vitamin D target cytochrome P450, family 24, subfamily A, polypeptide 1 (CYP24A1), demonstrating the presence of an autoregulatory vitamin D-endocrine system in these cells. Luciferase reporter studies demonstrated that cytochrome P450, family 27, subfamily B, polypeptide 1 (CYP27B1) was functional in these cells. Both 25OHD and 1,25(OH)(2)D altered C2C12 proliferation and differentiation. These effects were related to the increased expression of genes involved in G(0)/G(1) arrest (retinoblastoma protein [Rb], 1.3-fold; ATM, 1.5-fold, both P < .05), downregulation of mRNAs involved in G(1)/S transition, including myc and cyclin-D1 (0.7- and 0.8-fold, both P < .05) and reduced phosphorylation of Rb protein (0.3-fold, P < .005). After serum depletion, 1,25(OH)(2)D (100nM) suppressed myotube formation with decreased mRNAs for key myogenic regulatory factors (myogenin, 0.5-fold; myf5, 0.4-fold, P < .005) but led to a 1.8-fold increase in cross-sectional size of individual myotubes associated with markedly decreased myostatin expression (0.2-fold, P < .005). These data show that vitamin D signaling alters gene expression in C2C12 cells, with effects on proliferation, differentiation, and myotube size.

Beta-cell ARNT is required for normal glucose tolerance in murine pregnancy.

AIMS: Insulin secretion increases in normal pregnancy to meet increasing demands. Inability to increase beta-cell function results in gestational diabetes mellitus (GDM). We have previously shown that the expression of the transcription factor ARNT (Aryl-hydrocarbon Receptor Nuclear Translocator) is reduced in the islets of humans with type 2 diabetes. Mice with a beta-cell specific deletion of ARNT (ß-ARNT mice) have impaired glucose tolerance secondary to defective insulin secretion. We hypothesised that ARNT is required to increase beta-cell function during pregnancy, and that ß-ARNT mice would be unable to compensate for the beta-cell stress of pregnancy. The aims of this study were to investigate the mechanisms of ARNT regulation of beta-cell function and glucose tolerance in pregnancy. METHODS: ß-ARNT females were mated with floxed control (FC) males and FC females with ß-ARNT males. RESULTS: During pregnancy, ß-ARNT mice had a marked deterioration in glucose tolerance secondary to defective insulin secretion. There was impaired beta-cell proliferation in late pregnancy, associated with decreased protein and mRNA levels of the islet cell-cycle regulator cyclinD2. There was also reduced expression of Irs2 and G6PI. In contrast, in control mice, pregnancy was associated with a 2.1-fold increase in ARNT protein and a 1.6-fold increase in cyclinD2 protein, and with increased beta-cell proliferation. CONCLUSIONS: Islet ARNT increases in normal murine pregnancy and beta-cell ARNT is required for cyclinD2 induction and increased beta-cell proliferation in pregnancy.

Hypoxia-inducible factor-1α (HIF-1α) potentiates ß-cell survival after islet transplantation of human and mouse islets.

A high proportion of ß-cells die within days of islet transplantation. Reports suggest that induction of hypoxia-inducible factor-1α (HIF-1α) predicts adverse transplant outcomes. We hypothesized that this was a compensatory response and that HIF-1α protects ß-cells during transplantation. Transplants were performed using human islets or murine ß-cell-specific HIF-1α-null (ß-HIF-1α-null) islets with or without treatment with deferoxamine (DFO) to increase HIF-1α. ß-HIF-1α-null transplants had poor outcomes, demonstrating that lack of HIF-1α impaired transplant efficiency. Increasing HIF-1α improved outcomes for mouse and human islets. No effect was seen in ß-HIF-1α-null islets. The mechanism was decreased apoptosis, resulting in increased ß-cell mass posttransplantation. These findings show that HIF-1α is a protective factor and is required for successful islet transplant outcomes. Iron chelation with DFO markedly improved transplant success in a HIF-1α-dependent manner, thus demonstrating the mechanism of action. DFO, approved for human use, may have a therapeutic role in the setting of human islet transplantation.

High passage MIN6 cells have impaired insulin secretion with impaired glucose and lipid oxidation.

Type 2 diabetes is a metabolic disorder characterized by the inability of beta-cells to secrete enough insulin to maintain glucose homeostasis. MIN6 cells secrete insulin in response to glucose and other secretagogues, but high passage (HP) MIN6 cells lose their ability to secrete insulin in response to glucose. We hypothesized that metabolism of glucose and lipids were defective in HP MIN6 cells causing impaired glucose stimulated insulin secretion (GSIS). HP MIN6 cells had no first phase and impaired second phase GSIS indicative of global functional impairment. This was coupled with a markedly reduced ATP content at basal and glucose stimulated states. Glucose uptake and oxidation were higher at basal glucose but ATP content failed to increase with glucose. HP MIN6 cells had decreased basal lipid oxidation. This was accompanied by reduced expressions of Glut1, Gck, Pfk, Srebp1c, Ucp2, Sirt3, Nampt. MIN6 cells represent an important model of beta cells which, as passage numbers increased lost first phase but retained partial second phase GSIS, similar to patients early in type 2 diabetes onset. We believe a number of gene expression changes occurred to produce this defect, with emphasis on Sirt3 and Nampt, two genes that have been implicated in maintenance of glucose homeostasis.

Novel links between HIFs, type 2 diabetes, and metabolic syndrome.

Hypoxia inducible factors (HIFs) are master-regulators of cellular responses to hypoxia, and thus are crucial for survival. HIFs also play a role in regulating cellular processes in ß-cells, liver, muscle, and adipose tissue, have effects on the regulation of weight, and play a role in type 2 diabetes (T2D). Indeed, in people with T2D the HIF pathway is dyregulated in major metabolic tissues involved in the pathogenesis of diabetes. This review covers the contrasting, complementary and conflicting effects of decreasing and increasing HIFs in various tissues, and shows that a delicate balance exists between HIF levels and optimal metabolic function. We propose that increasing the activity of HIFs might be a potential therapeutic strategy for treating T2D.

Mice deficient in GEM GTPase show abnormal glucose homeostasis due to defects in beta-cell calcium handling.

AIMS AND HYPOTHESIS: Glucose-stimulated insulin secretion from beta-cells is a tightly regulated process that requires calcium flux to trigger exocytosis of insulin-containing vesicles. Regulation of calcium handling in beta-cells remains incompletely understood. Gem, a member of the RGK (Rad/Gem/Kir) family regulates calcium channel handling in other cell types, and Gem over-expression inhibits insulin release in insulin-secreting Min6 cells. The aim of this study was to explore the role of Gem in insulin secretion. We hypothesised that Gem may regulate insulin secretion and thus affect glucose tolerance in vivo. METHODS: Gem-deficient mice were generated and their metabolic phenotype characterised by in vivo testing of glucose tolerance, insulin tolerance and insulin secretion. Calcium flux was measured in isolated islets. RESULTS: Gem-deficient mice were glucose intolerant and had impaired glucose stimulated insulin secretion. Furthermore, the islets of Gem-deficient mice exhibited decreased free calcium responses to glucose and the calcium oscillations seen upon glucose stimulation were smaller in amplitude and had a reduced frequency. CONCLUSIONS: These results suggest that Gem plays an important role in normal beta-cell function by regulation of calcium signalling.

Effects of environmentally relevant concentrations of endosulfan, azinphosmethyl, and diazinon on Great Basin spadefoot (Spea intermontana) and Pacific treefrog (Pseudacris regilla).

We conducted dose-response exposures to compare the lethality of endosulfan, diazinon, and azinphosmethyl in the early-life stages of the Great Basin spadefoot (Spea intermontana) and the Pacific treefrog (Pseudacris regilla). Our experiment occurred in two 8-d phases: one, with developing embryos, and two, with Gosner Stage 27 tadpoles. Pesticide concentrations were representative of field-measured concentrations (60 ng/L of endosulfan, 50 ng/L of azinphosmethyl, and 350 ng/L of diazinon), in the same geographic areas where these species occur in British Columbia. Although the concentrations met the requirements for federal water quality guidelines, we observed mortalities, deformities, and other sublethal effects. Phase 1 consisted of exposing Gosner Stage 10 embryos in the pesticide solutions for a total of 8 d. Significant mortality of S. intermontana began posthatch in the highest lethal concentrations of the commercial formulations of endosulfan (Thiodan; LC20(8d)=2,672.7 ng/L) and diazinon (LC20(8d)>175,000 ng/L). Phase 2 compared behavior, morphology, and survival of captive-reared tadpoles exposed to the same 8-d experimental regime as the embryo experiment. Endosulfan induced significant effects on behavior and morphology of P. regilla and significantly reduced survivorship of S. intermontana (LC20(8d)=77.1 ng/L). Abnormal behavior and excitability was observed in both species, with P. regilla tadpoles being more sensitive. At 60,000 ng/L endosulfan, P. regilla also lost pigmentation and exhibited abnormal tail morphology.

Hypoxia-inducible factor-1alpha regulates beta cell function in mouse and human islets.

Hypoxia-inducible factor-1alpha (HIF-1alpha) is a transcription factor that regulates cellular stress responses. While the levels of HIF-1alpha protein are tightly regulated, recent studies suggest that it can be active under normoxic conditions. We hypothesized that HIF-1alpha is required for normal beta cell function and reserve and that dysregulation may contribute to the pathogenesis of type 2 diabetes (T2D). Here we show that HIF-1alpha protein is present at low levels in mouse and human normoxic beta cells and islets. Decreased levels of HIF-1alpha impaired glucose-stimulated ATP generation and beta cell function. C57BL/6 mice with beta cell-specific Hif1a disruption (referred to herein as beta-Hif1a-null mice) exhibited glucose intolerance, beta cell dysfunction, and developed severe glucose intolerance on a high-fat diet. Increasing HIF-1alpha levels by inhibiting its degradation through iron chelation markedly improved insulin secretion and glucose tolerance in control mice fed a high-fat diet but not in beta-Hif1a-null mice. Increasing HIF-1alpha levels markedly increased expression of ARNT and other genes in human T2D islets and improved their function. Further analysis indicated that HIF-1alpha was bound to the Arnt promoter in a mouse beta cell line, suggesting direct regulation. Taken together, these findings suggest an important role for HIF-1alpha in beta cell reserve and regulation of ARNT expression and demonstrate that HIF-1alpha is a potential therapeutic target for the beta cell dysfunction of T2D.

Oxygen initiation of respiration and mitochondrial biogenesis in rice.

Rice growth under aerobic and anaerobic conditions allowed aspects of mitochondrial biogenesis to be identified as dependent on or independent of an oxygen signal. Analysis of transcripts encoding mitochondrial components found that a subset of these genes respond to oxygen (defined as aerobic), whereas others are relatively unaffected by oxygen availability. Mitochondria formed during growth in anaerobic conditions had reduced protein levels of tricarboxylic acid cycle components and cytochrome-containing complexes of the respiratory chain and repressed respiratory functionality. In general, the capacity of the general import pathway was found to be significantly lower in mitochondria isolated from tissue grown under anaerobic conditions, whereas the carrier import pathway capacity was not affected by changes in oxygen availability. Transcript levels of genes encoding components of the protein import apparatus were generally not affected by the absence of oxygen, and their protein abundance was severalfold higher in mitochondria isolated from anaerobically grown tissue. However, both transcript and protein abundances of the subunits of the mitochondrial processing peptidase, which in plants is integrated into the cytochrome bc(1) complex, were repressed under anaerobic conditions. Therefore, in this system, an increase in import capacity is correlated with an increase in the abundance of the cytochrome bc(1) complex, which is ultimately dependent on the presence of oxygen, providing a link between the respiratory chain and protein import apparatus.