A C2H2 Zinc Finger Protein FEMU2 Is Required for fox1 Expression in Chlamydomonas reinhardtii

Abstract

Chlamydomonas reinhardtii fox1 gene encodes a ferroxidase that is involved in cellular Fe uptake and highly induced during Fe deficient conditions. In an effort to identify fox1 promoter regulatory elements, an insertional library was generated in a transgenic Chlamydomonas strain (2A38) harboring an arylsulfatase (ARS) reporter gene driven by the fox1 promoter. Mutants with a defective response to low iron conditions were selected for further study. Among these, a strain containing a disrupted femu2 gene was identified. Activation of the fox1 promoter by the femu2 gene product was confirmed by silencing the femu2 gene using RNA interference. In three femu2 RNAi transgenic lines (IR3, IR6, and IR7), ARS reporter gene activities declined by 84.3%, 86.4%, and 88.8%, respectively under Fe deficient conditions. Furthermore, RT-PCR analysis of both the femu2 mutant and the RNAi transgenic lines showed significantly decreased transcript abundance of the endogenous fox1 gene under Fe deficient conditions. Amino acid sequence analysis of the femu2 gene product identified three potential C2H2 zinc finger (ZF) motifs and a nuclear localization study suggests that FEMU2 is localized to the nucleus. In addition, a potential FEMU2 binding site ((G/T)TTGG(G/T)(G/T)T) was identified using PCR-mediated random binding site selection. Taken together, this evidence suggests that FEMU2 is involved in up-regulation of the fox1 gene in Fe deficient cells.

Diversity, expansion, and evolutionary novelty of plant DNA-binding transcription factor families

“…The regulation of genes in response to communication between individual cells and subsequent signaling cascades is enabled by common TFs such as Homeobox [93] and MADS-box [28]. In addition, Heat Shock Factor (HSF, [94]) and Zinc finger (Zf, [95]) TFs are important regulatory components of cellular stress response, and bZIP family members are important for the endoplasmic reticulum stress response [96]. Although TF families that are found in most eukaryotes are expected to be important for cellular functions, they have often gained novel regulatory roles.…”

“…Although TF families that are found in most eukaryotes are expected to be important for cellular functions, they have often gained novel regulatory roles. For example, cell fate determination, cellular differentiation, and/or multiple stages of multicellular eukaryotic development are regulated by several common TF families including Myb [97], bHLH [76], bZIP [98], ARID [77], and C2H2 Zf [95]. Myb [74] and CBFB/NF-Y [99] also regulate apoptosis and cell senescence.…”

Regulating cellular trace metal economy in algae

“…Proteins potentially involved in FOX1 expression have also been identified through a genetic screen assaying absence of FOX1 -promoter-arylsulfatase fusion activity during Fe starvation. The corresponding loci encode a putative protein kinase [63] and a RING-type Zn finger protein that contains a Ran-binding domain (Ran regulates receptor-mediated transport between the nucleus and cytoplasm) [64]. However, it remains unknown whether these two proteins are directly involved in Fe-responsive signaling.…”