Cloning of a full‐length symbiotic hemoglobin cDNA and in situ localization of the corresponding mRNA in Casuarina glauca root nodule

Abstract

gusz, D, 1997, Cloning of a full-length symbiotic hemoglobin cDNA and in situ localization of the corresponding mRNA in Casuarina giauca root nodule, -Physiol, Plant, 99:608-616,We have characterized a full-length cDNA (hb-Cg\¥) that represents symbiotic mRNA hemoglobin (hb) from Casuarina glauca root nodules. In situ hybridization was used to examine the correlation between hb-CgW mRNA and the state of the Frankia infection process. The efficiency of in situ hybridization using DIG-labeled vs ['^SJ-labeled probes was compared. The expression of hb-CgW gene is induced in young infected host cells prior to the detection of Frankia nifH mRNA, Since Frankia does not form vesicles in C, giauca nodules, it is proposed that Hb is necessary to reduce the O; concentration in the cytoplasm of the host cells before the nif genes are expressed.

Transcriptomics of Actinorhizal Symbioses Reveals Homologs of the Whole Common Symbiotic Signaling Cascade

“…Alnus glutinosa seeds were harvested from an A. glutinosa specimen growing on the left bank of the Rhô ne River in Lyon, France, used previously (Alloisio et al, 2010). Both species were grown and inoculated with a compatible Frankia strain: CcI3 for C. glauca (Gherbi et al, 1997) and ACN14a for A. glutinosa (Normand and Lalonde, 1982), as described previously (Gherbi et al, 1997;Alloisio et al, 2010). For construction of the EST library, noninoculated roots (controls), inoculated roots (2 and 7 d dpi), and nodules (3 weeks post inoculation) were collected and immediately frozen in liquid nitrogen.…”

“…In Alnus, the oxygen level is not regulated, and Frankia has to adapt by synthesizing specialized cells called vesicles with thick hopanoid walls that form a diffusion barrier (Berry et al, 1993). When vesicles are absent, as in Casuarina, there is a low-oxygen tension in the infected cells (Tjepkema, 1979), presumably due to the lignification of their cell walls (Berg and McDowell, 1988), whereas a symbiotic hemoglobin facilitates the supply of oxygen to the bacterial respiration chain (Gherbi et al, 1997). In addition, nodular roots are prevalent in Casuarina and very rare in Alnus (Torrey, 1976); these peculiar roots, which emerge from the nodule apex, show a negative geotropism, are free of root hair and of bacterial infection, and are thought to permit the supply of oxygen and nitrogen to nodules under water saturation conditions (Tjepkema, 1978).…”

Characterization of a Casuarina glauca Nodule-Specific Subtilisin-like Protease Gene, a Homolog of Alnus glutinosa ag12

“…cg12 probes were prepared by linearization of pCg12 with SstII and in vitro transcription with T7 RNA polymerase (antisense) or linearization with PstI and in vitro transcription with T3 RNA polymerase (sense), respectively. Parallel sections were hybridized with a probe for the nitrogenase structural gene nifH from Frankia, prepared as described by Gherbi et al (1997) to identify infected cells containing nitrogen-fixing bacteria. cg12 hybridization signals were strong in young, infected cells of the infection zone and weaker and irregular in the infected cells of the fixation zone as denoted by Frankia nifH expression (Fig.…”

Casuarina glauca Prenodule Cells Display the Same Differentiation as the Corresponding Nodule Cells

“…Expression of nifH, encoding a subunit of the nitrogenase complex, denotes bacterial differentiation for symbiotic nitrogen fixation (Berg and McDowell 1987a;Ribeiro et al 1995). In C. glauca nodules, nitrogen-fixing infected cells differentiate to provide (i) an oxygen diffusion barrier achieved by lignification of the cell wall (Berg and McDowell 1987b) and (ii) a high amount of the oxygen-transport protein hemoglobin (Jacobsen-Lyon et al 1995;Gherbi et al 1997). Frankia nifH and C. glauca symbiotic hemoglobin (hb) gene expression was examined by in situ hybridization.…”

“…nifH and hb probes were prepared as described by Gherbi et al (1997). cg12 probes were prepared by linearization of pCg12 with SstII and in vitro transcription with T7 RNA polymerase (antisense) or by linearization with PstI and transcription with T3 RNA polymerase (sense).…”