Rac1 Dynamics in the Human Opportunistic Fungal Pathogen Candida albicans

The small Rho G-protein Rac1 is highly conserved from fungi to humans, with approximately 65% overall sequence identity in Candida albicans. As observed with human Rac1, we show that C. albicans Rac1 can accumulate in the nucleus, and fluorescence recovery after photobleaching (FRAP) together with fluorescence loss in photobleaching (FLIP) studies indicate that this Rho G-protein undergoes nucleo-cytoplasmic shuttling. Analyses of different chimeras revealed that nuclear accumulation of C. albicans Rac1 requires the NLS-motifs at its carboxyl-terminus, which are blocked by prenylation of the adjacent cysteine residue. Furthermore, we show that C. albicans Rac1 dynamics, both at the plasma membrane and in the nucleus, are dependent on its activation state and in particular that the inactive form accumulates faster in the nucleus. Heterologous expression of human Rac1 in C. albicans also results in nuclear accumulation, yet accumulation is more rapid than that of C. albicans Rac1. Taken together our results indicate that Rac1 nuclear accumulation is an inherent property of this G-protein and suggest that the requirements for its nucleo-cytoplasmic shuttling are conserved from fungi to humans

Phenotypic characterization of breast cancer: the role of CDC42

Purpose: The molecular landscape of breast cancer (BC), especially of the Luminal A subtype, remains to be fully delineated. Transcriptomic data shows that Luminal A tumours are enriched for aberrant expression of genes in the cell division control 42 homolog (CDC42) pathway. This study aims to investigate protein expression of CDC42 in BC and assess its clinicopathological significance. Methods: Expression of CDC42 protein was examined by immunohistochemistry on tissue microarrays in a well characterised cohort of 895 early stage (I-IIIa) primary invasive BCs. Results: CDC42 expression was observed in both the cytoplasm and nucleus of BC cells. High nuclear CDC42 expression demonstrated a significant correlation with ER positive, low-grade tumours and was more common in the lobular histological subtype (all p<0.001). In contrast, cytoplasmic CDC42 showed increased expression in the ductal subtype (p<0.001) and correlated with negative prognostic features such as larger size, higher grade (p<0.05), and higher Ki67 labelling index (p=0.001). Nuclear CDC42 expression was associated with a longer BC specific survival in all cases (p=0.025) and in luminal ER positive tumours (p=0.011). In multivariate analyses including size, grade, lymph node stage and intrinsic subtype, CDC42 was an independent prognostic factor (p=0.032). Conclusion: The results indicate that CDC42 is important molecule in luminal BC, with prognostic significance

Alcohol-derived DNA crosslinks are repaired by two distinct mechanisms

Acetaldehyde is a highly reactive, DNA-damaging metabolite that is produced upon alcohol consumption1. Impaired detoxification of acetaldehyde is common in the Asian population, and is associated with alcohol-related cancers1,2. Cells are protected against acetaldehyde-induced damage by DNA crosslink repair, which when impaired causes Fanconi anaemia (FA), a disease resulting in failure to produce blood cells and a predisposition to cancer3,4. The combined inactivation of acetaldehyde detoxification and the FA pathway induces mutation, accelerates malignancies and causes the rapid attrition of blood stem cells5,6,7. However, the nature of the DNA damage induced by acetaldehyde and how this is repaired remains a key question. Here we generate acetaldehyde-induced DNA interstrand crosslinks and determine their repair mechanism in Xenopus egg extracts. We find that two replication-coupled pathways repair these lesions. The first is the FA pathway, which operates using excision—analogous to the mechanism used to repair the interstrand crosslinks caused by the chemotherapeutic agent cisplatin. However, the repair of acetaldehyde-induced crosslinks results in increased mutation frequency and an altered mutational spectrum compared with the repair of cisplatin-induced crosslinks. The second repair mechanism requires replication fork convergence, but does not involve DNA incisions—instead the acetaldehyde crosslink itself is broken. The Y-family DNA polymerase REV1 completes repair of the crosslink, culminating in a distinct mutational spectrum. These results define the repair pathways of DNA interstrand crosslinks caused by an endogenous and alcohol-derived metabolite, and identify an excision-independent mechanism

A survey of the ATP-binding cassette (ABC) gene superfamily in the salmon louse (Lepeophtheirus salmonis)

Salmon lice,Lepeophtheirus salmonis(Kr&oslash;yer, 1837), are fish ectoparasites causing significant economic damage in the mariculture of Atlantic salmon,Salmo salarLinnaeus, 1758. The control ofL.salmonisat fish farms relies to a large extent on treatment with anti-parasitic drugs. A problem related to chemical control is the potential for development of resistance, which inL.salmonisis documented for a number of drug classes including organophosphates, pyrethroids and avermectins. The ATP-binding cassette (ABC) gene superfamily is found in all biota and includes a range of drug efflux transporters that can confer drug resistance to cancers and pathogens. Furthermore, some ABC transporters are recognised to be involved in conferral of insecticide resistance. While a number of studies have investigated ABC transporters inL.salmonis, no systematic analysis of the ABC gene family exists for this species. This study presents a genome-wide survey of ABC genes inL.salmonisfor which, ABC superfamily members were identified through homology searching of theL.salmonisgenome. In addition, ABC proteins were identified in a reference transcriptome of the parasite generated by high-throughput RNA sequencing (RNA-seq) of a multi-stage RNA library. Searches of both genome and transcriptome allowed the identification of a total of 33 genes / transcripts coding for ABC proteins, of which 3 were represented only in the genome and 4 only in the transcriptome. Eighteen sequences were assigned to ABC subfamilies known to contain drug transporters,i.e. subfamilies B (4 sequences), C (11) and G (2). The results suggest that the ABC gene family ofL.salmonispossesses fewer members than recorded for other arthropods. The present survey of theL.salmonisABC gene superfamily will provide the basis for further research into potential roles of ABC transporters in the toxicity of salmon delousing agents and as potential mechanisms of drug resistance

Bed bugs evolved unique adaptive strategy to resist pyrethroid insecticides

Recent advances in genomic and post-genomic technologies have facilitated a genome-wide analysis of the insecticide resistance-associated genes in insects. Through bed bug, Cimex lectularius transcriptome analysis, we identified 14 molecular markers associated with pyrethroid resistance. Our studies revealed that most of the resistance-associated genes functioning in diverse mechanisms are expressed in the epidermal layer of the integument, which could prevent or slow down the toxin from reaching the target sites on nerve cells, where an additional layer of resistance (kdr) is possible. This strategy evolved in bed bugs is based on their unique morphological, physiological and behavioral characteristics and has not been reported in any other insect species. RNA interference-aided knockdown of resistance associated genes showed the relative contribution of each mechanism towards overall resistance development. Understanding the complexity of adaptive strategies employed by bed bugs will help in designing the most effective and sustainable bed bug control methods