The MADS-box gene XAANTAL1 participates in Arabidopsis thaliana primary root growth and columella stem cell patterns in response to ROS, via direct regulation of PEROXIDASE 28 and RETINOBLASTOMA RELATED.

The balance between cell growth, proliferation and differentiation emerges from gene regulatory networks coupled to various signal transduction pathways, including reactive oxygen species (ROS) and transcription factors (TFs), enabling developmental responses to environmental cues. The Arabidopsis thaliana's primary root has become a valuable system for unraveling such networks. Recently, the role of TFs that mediate the ROS's inhibition of primary root growth has begun to be characterized. This study demonstrates that the MADS-box transcription factor XAANTAL1 (XAL1) is an essential regulator of hydrogen peroxide (H2O2) in primary root growth and root stem cell niche identity. Interestingly, our findings suggest that XAL1 acts as a positive regulator of H2O2 concentration in the root meristem by directly regulating genes involved in oxidative stress response, such as PEROXIDASE 28 (PER28). Moreover, we found that XAL1 is necessary for the H2O2-induced inhibition of primary root growth through the negative regulation of peroxidase and catalase activities. Furthermore, XAL1, in conjunction with RETINOBLASTOMA-RELATED (RBR), is essential for positively regulating the differentiation of columella stem cells and for participating in primary root growth inhibition in response to oxidative stress induced by H2O2 treatment.

The effect of iron on Trichomonas vaginalis TvCP2: a cysteine proteinase found in vaginal secretions of trichomoniasis patients.

Trichomonas vaginalis (Tv) induces host cell damage through cysteine proteinases (CPs) modulated by iron. An immunoproteomic analysis showed that trichomoniasis patient sera recognize various CPs, also some of them are present in vaginal washes (VWs). Thus, the goal of this work was to determine whether TvCP2 is expressed during infection and to assess the effect of iron on TvCP2 expression, localization and contribution to in vitro cellular damage. Western-blotting (WB) assays using TvCP2r and vaginitis patient serum samples showed that 6/9 Tv (+) but none of the Tv (-) patient sera recognized TvCP2r. WB using an anti-TvCP2r antibody and VWs from the same patients showed that in all of the Tv (+) but none of the Tv (-) VWs, the anti-TvCP2r antibody detected a 27 kDa protein band that corresponded to the mature TvCP2, which was confirmed by mass spectrometry analysis. Iron decreased the amount of TvCP2 mRNA and the protein localized on the parasite surface and cytoplasmic vesicles concomitant with the cytotoxic effect of TvCP2 on HeLa cells. Parasites pretreated with the anti-TvCP2r antibody also showed reduced levels of cytotoxicity and apoptosis induction in HeLa cell monolayers. In conclusion, these results show that TvCP2 is expressed during trichomonal infection and plays an important role in the in vitro HeLa cell cytotoxic damage under iron-restricted conditions.

The MADS-box genes SOC1 and AGL24 antagonize XAL2 functions in Arabidopsis thaliana root development.

MADS-domain transcription factors play pivotal roles in numerous developmental processes in Arabidopsis thaliana. While their involvement in flowering transition and floral development has been extensively examined, their functions in root development remain relatively unexplored. Here, we explored the function and genetic interaction of three MADS-box genes (XAL2, SOC1 and AGL24) in primary root development. By analyzing loss-of-function and overexpression lines, we found that SOC1 and AGL24, both critical components in flowering transition, redundantly act as repressors of primary root growth as the loss of function of either SOC1 or AGL24 partially recovers the primary root growth, meristem cell number, cell production rate, and the length of fully elongated cells of the short-root mutant xal2-2. Furthermore, we observed that the simultaneous overexpression of AGL24 and SOC1 leads to short-root phenotypes, affecting meristem cell number and fully elongated cell size, whereas SOC1 overexpression is sufficient to affect columella stem cell differentiation. Additionally, qPCR analyses revealed that these genes exhibit distinct modes of transcriptional regulation in roots compared to what has been previously reported for aerial tissues. We identified 100 differentially expressed genes in xal2-2 roots by RNA-seq. Moreover, our findings revealed that the expression of certain genes involved in cell differentiation, as well as stress responses, which are either upregulated or downregulated in the xal2-2 mutant, reverted to WT levels in the absence of SOC1 or AGL24.

Characterization of a novel endogenous cysteine proteinase inhibitor, trichocystatin-3 (TC-3), localized on the surface of Trichomonas vaginalis.

Trichomonas vaginalis is a flagellated protist responsible for human trichomoniasis. T. vaginalis has three genes encoding for endogenous cysteine proteinase (CP) inhibitors, known as trichocystatin-1 through trichocystatin-3 (TC-1, TC-2, and TC-3). These inhibitors belong to the cystatin family. In this study, we characterized trichocystatin-3 (TC-3), an endogenous cysteine proteinase (CP) inhibitor of T. vaginalis. TC-3 possesses a signal peptide in the N-terminus and two putative glycosylation sites (typical of family 2, cystatins) but lacks the PW motif and cysteine residues (typical of family 1, stefins). Native TC-3 was recognized as an ∼18 kDa protein band in a T. vaginalis protein extract. By confocal microscopy, endogenous TC-3 was found in the Golgi complex, cytoplasm, large vesicles, and the plasma membrane. These localizations are consistent with an in silico prediction. In addition, the purified recombinant protein (TC-3r) functions as an inhibitor of cathepsin L CPs, such as human liver cathepsin L and trichomonad CPs, present in a proteinase-resistant extract (PRE). Via a pull-down assay using TC-3r as bait and PRE, we identified several trichomonad CPs targeted by TC-3, primarily TvCP3. These CP-TC-3 interactions occur in vesicles, in the cytoplasm, and on the parasite surface. In addition, TC-3r showed a protective effect on HeLa cell monolayers by inhibiting trichomonad surface CPs involved in cellular damage. Our results show that the endogenous inhibitor TC-3 plays a key role in the regulation of endogenous CP proteolytic activity.