Infection thread-dependent invasion of legume roots by rhizobia leads to internalization of bacteria into the plant cells, which is one of the salient features of root nodule symbiosis. We found that two genes, Nap1 (for Nck-associated protein 1) and Pir1 (for 121F-specific p53 inducible RNA), involved in actin rearrangements were essential for infection thread formation and colonization of Lotus japonicus roots by its natural microsymbiont, Mesorhizobium loti. nap1 and pir1 mutants developed an excess of uncolonized nodule primordia, indicating that these two genes were not essential for the initiation of nodule organogenesis per se. However, both the formation and subsequent progression of infection threads into the root cortex were significantly impaired in these mutants. We demonstrate that these infection defects were due to disturbed actin cytoskeleton organization. Short root hairs of the mutants had mostly transverse or web-like actin filaments, while bundles of actin filaments in wild-type root hairs were predominantly longitudinal. Corroborating these observations, temporal and spatial differences in actin filament organization between wild-type and mutant root hairs were also observed after Nod factor treatment, while calcium influx and spiking appeared unperturbed. Together with various effects on plant growth and seed formation, the nap1 and pir1 alleles also conferred a characteristic distorted trichome phenotype, suggesting a more general role for Nap1 and Pir1 in processes establishing cell polarity or polar growth in L. japonicus.

Sandal et al. MPMI 4 INTRODUCTIONGenetic analysis and application of genetic approaches in the model legume Lotus japonicus (Handberg and Stougaard 1992) has progressed rapidly. Several key genes important for symbiosis with mycorrhizal fungi, root nodule development and other developmental processes have been identified using molecular genetics. The developmental regulators Nin (Schauser et al. 1999) and Pfo (Zhang et al. 2002) were isolated by transposon tagging while map-based cloning led to the molecular characterisation of Har1, SymRK, Nfr1, Nfr5, Castor and Pollux involved in autoregulation, Nod-factor signal perception or signal transduction (Schauser et al. 1999, Krusell et al. 2002 Nishimura et al. 2002a;Stracke et al. 2002;Radutoiu et al. 2003;Madsen et al. 2003; Imaizumi-Anraku et al. 2005). Genetic loci required for the early stages of endosymbiosis have attracted particular interest. Diallelic crosses together with phenotypical studies defined seven loci, SymRK, Nup133, Castor, Pollux, Sym6, Sym15,Sym24, in the common pathway required for both rhizobial and mycorrhizal symbiosis (Kistner et al. unpublished data) and map-based cloning of these loci has been accomplished or is advancing rapidly. A similar interest and effort is now emerging for genetic dissection of nodule organogenesis and function using the Fix -mutants arrested at various stages of nodule development or impaired in nodule function. Cloning of the Sst1 sulfate transporter required in functional root nodules is a first example (Krusell et al. 2005).Continuous isolation of new plant mutant lines is important for completing the genetic dissection of symbiosis and so far six independent mutant populations have been obtained by chemical (EMS) mutagenesis (Perry et al. 2003;Szczyglowski et al. 1998; Webb et al. unpublished data; Gresshoff et al. unpublished data), four populations after T-DNA or transposon insertion mutagenesis (Thykjaer et al. 1995;Schauser et al. 1998;Webb et al. 2000; Gresshoff et al. unpublished data), one population made with fast neutrons (Gresshoff et al. unpublished Umehara and Kouchi (unpublished data). All in all more than 400 symbiotic Lotus mutant lines were identified by screening in these populations and more are likely to follow. Assignment to complementation groups is next logical step in order to determine the number of loci involved, identify all alleles that contribute to phenotypic characterisation of mutants and genotyping of loci. However, diallelic crossing is a relatively slow process where progress is determined by generation time and slowed by a continuously increasing number of individual crosses necessary to keep up with mutant isolation programs. Given the number of symbiotic mutant lines already available and considering the time used to define seven complementation groups with a total of 26 alleles constituting the common pathway (Kistner et al. unpublished data), this approach is unlikely to encompass all alleles in near future. Detection of alleles in already cloned genes ...

Abstract. Glucose is the major source of energy for cells, and glucose transporter 1 (GLUT1) is the most common glucose transporter. GLUT1 has been found to be aberrantly expressed in several tumor types. From the results of the microarray and serial analysis of gene expression (SAGE), GLUT1 transcript expression was found to be higher in clones with mutant Kras alleles. We hypothesized that GLUT1 overexpression might be correlated with clinicopathological features of Japanese lung cancers. Immunohistochemistry for GLUT1 was performed in 283 surgically treated non-small cell lung cancer (NSCLC) cases from Nagoya City University Hospital. Thirty-six Kras mutant carcinoma cases were included. GLUT1 overexpression was found in 138 (48.8%) lung cancer patients. The GLUT1 overexpression status was significantly correlated with gender (women 31.9% vs. men 54.5%, P<0.0001), smoking status (never smoker 31.4% vs. smoker 59.4%, P<0.0001) and pathological subtypes (adenocarcinoma 36.4% vs. non-adenocarcinoma 74.5%, P<0.0001). In addition, the GLUT1 overexpression status was significantly correlated with gene mutation status, including EGFR (mutation-positive 23.4% vs. -negative 58.3%, P<0.0001) and Kras (mutation-positive 66.7% vs. -negative 46.6%, P=0.038). The survival of patients with GLUT1 overexpression (n=137, 50 were deceased) was significantly worse when compared to the patients with normal expression of GLUT1 (n=142, 31 were deceased) (Log-rank test, P=0.0009). Thus, GLUT-1 overexpression correlates with an aggressive phenotype of lung carcinoma.