Macroautophagy/autophagy plays a dual role in many physiological processes of multicellular eukaryotes. In plants, autophagy can be used by both host and pathogen for a beneficiary infection outcome. Plants employ a two-tier innate immune system to defend against invading pathogens. Cell surface localized pattern recognition receptors recognize conserved pathogen-associated molecular patterns (PAMPs) and launch pattern-triggered immunity (PTI) to provide broad-spectrum resistance. Pathogens inject a battery of effector proteins into their hosts to counter PTI and compromise the primary immune response. Hosts induce a second layer of defense called effector-triggered immunity (ETI) to counter the effects of these effectors. In addition to ETI and PTI, autophagy is emerging as a central cellular process modulated by both host and pathogens toward their respective advantage. Pathogens lacking the ability to inject effectors are compromised in virulence. However, molecular targets and biochemical characterization of most of these effector proteins remain elusive. In a recent paper we presented a systematic analysis of interaction between autophagy proteins of Arabidopsis thaliana with effectors from bacterial, fungal, oomycete and nematode pathogens. Abbreviations: ATG, autophagy related; BiFC, bimolecular fluorescence complementation; ETI, effector-triggered immunity; PAMPs, pathogen-associated molecular patterns; PTI, pattern-triggered immunity.

Spodoptera frugiperda (fall armyworm) is a notorious pest that threatens maize production worldwide. Current control measures involve the use of chemical insecticides and transgenic maize expressing Bacillus thuringiensis (Bt) toxins. Although additional transgenes have confirmed insecticidal activity, limited research has been conducted in maize, at least partially due to the technical difficulty of maize transformation. Here, we describe implementation of a sugarcane mosaic virus (SCMV) vector for rapidly testing the efficacy of both endogenous maize genes and heterologous genes from other organisms for the control of S. frugiperda in maize. Four categories of proteins were tested using the SCMV vector: (i) maize defence signalling proteins: peptide elicitors (Pep1 and Pep3) and jasmonate acid conjugating enzymes (JAR1a and JAR1b); (ii) maize defensive proteins: the previously identified ribosome-inactivating protein (RIP2) and maize proteinase inhibitor (MPI), and two proteins with predicted but unconfirmed anti-insect activities, an antimicrobial peptide (AMP) and a lectin (JAC1); (iii) lectins from other plant species: Allium cepa agglutinin (ACA) and Galanthus nivalis agglutinin (GNA); and (iv) scorpion and spider toxins: peptides from Urodacus yaschenkoi (UyCT3 and UyCT5) and Hadronyche versuta (Hvt). In most cases, S. frugiperda larval growth was reduced by transient SCMV-mediated overexpression of genes encoding these proteins. Additionally, experiments with a subset of the SCMV-expressed genes showed effectiveness against two aphid species, Rhopalosiphum maidis (corn leaf aphid) and Myzus persicae (green peach aphid). Together, these results demonstrate that SCMV vectors are a rapid screening method for testing the efficacy and insecticidal activity of candidate genes in maize.

Background

Polarity is necessary for epithelial cells to perform distinct functions at their apical and basal surfaces. Oral epithelial cell-derived ameloblasts at secretory stage (SABs) synthesize large amounts of enamel matrix proteins (EMPs), largely amelogenins. EMPs are unidirectionally secreted into the enamel space through their apical cytoplasmic protrusions, or Tomes' processes (TPs), to guide the enamel formation. Little is known about the transcriptional regulation underlying the establishment of cell polarity and unidirectional secretion of SABs.

Results

The higher-order chromatin architecture of eukaryotic genome plays important roles in cell- and stage-specific transcriptional programming. A genome organizer, special AT-rich sequence-binding protein 1 (SATB1), was discovered to be significantly upregulated in ameloblasts compared to oral epithelial cells using a whole-transcript microarray analysis. The Satb1^-/- mice possessed deformed ameloblasts and a thin layer of hypomineralized and non-prismatic enamel. Remarkably, Satb1^-/- ameloblasts at the secretory stage lost many morphological characteristics found at the apical surface of wild-type (wt) SABs, including the loss of Tomes' processes, defective inter-ameloblastic adhesion, and filamentous actin architecture. As expected, the secretory function of Satb1^-/- SABs was compromised as amelogenins were largely retained in cells. We found the expression of epidermal growth factor receptor pathway substrate 8 (Eps8), a known regulator for actin filament assembly and small intestinal epithelial cytoplasmic protrusion formation, to be SATB1 dependent. In contrast to wt SABs, EPS8 could not be detected at the apical surface of Satb1^-/- SABs. Eps8 expression was greatly reduced in small intestinal epithelial cells in Satb1^-/- mice as well, displaying defective intestinal microvilli.

Conclusions

Our data show that SATB1 is essential for establishing secretory ameloblast cell polarity and for EMP secretion. In line with the deformed apical architecture, amelogenin transport to the apical secretory front and secretion into enamel space were impeded in Satb1^-/- SABs resulting in a massive cytoplasmic accumulation of amelogenins and a thin layer of hypomineralized enamel. Our studies strongly suggest that SATB1-dependent Eps8 expression plays a critical role in cytoplasmic protrusion formation in both SABs and in small intestines. This study demonstrates the role of SATB1 in the regulation of amelogenesis and the potential application of SATB1 in ameloblast/enamel regeneration.