Crop plants contain large amounts of secondary compounds that interfere with protein extraction and gel-based proteomic analysis. Thus, a protein extraction protocol that can be easily applied to various crop materials with minimal optimization is essential. Here we describe a universal protocol for total protein extraction involving trichloroacetic acid (TCA)/acetone precipitation followed by SDS and phenol extraction. Through SDS extraction, the proteins precipitated by the TCA/acetone treatment can be fully resolubilized and then further purified by phenol extraction. This protocol combines TCA/acetone precipitation, which aggressively removes nonprotein compounds, and phenol extraction, which selectively dissolves proteins, resulting in effective purification of proteins from crop tissues. This protocol can also produce high-quality protein preparations from various recalcitrant tissues, and therefore it has a wide range of applications in crop proteomic analysis. Designed to run on a small scale, this protocol can be completed within 5 h.

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IgA is the most abundantly produced antibody in the body and plays a crucial role in gut homeostasis and mucosal immunity. IgA forms a dimer that covalently associates with the joining (J) chain, which is essential for IgA transport into the mucosa. Here, we demonstrate that the marginal zone B and B-1 cell-specific protein (MZB1) interacts with IgA through the α-heavy-chain tailpiece dependent on the penultimate cysteine residue and prevents the intracellular degradation of α-light-chain complexes. Moreover, MZB1 promotes J-chain binding to IgA and the secretion of dimeric IgA. MZB1-deficient mice are impaired in secreting large amounts of IgA into the gut in response to acute inflammation and develop severe colitis. Oral administration of a monoclonal IgA significantly ameliorated the colitis, accompanied by normalization of the gut microbiota composition. The present study identifies a molecular chaperone that promotes J-chain binding to IgA and reveals an important mechanism that controls the quantity, quality, and function of IgA.

Immunoglobulin (Ig) M is the first antibody isotype to appear during evolution, ontogeny and immune responses. IgM not only serves as the first line of host defense against infections but also plays an important role in immune regulation and immunological tolerance. For many years, IgM is thought to function by binding to antigen and activating complement system. With the discovery of the IgM Fc receptor (FcμR), it is now clear that IgM can also elicit its function through FcμR. In this review, we will describe the molecular characteristics of FcμR, its role in B cell development, maturation and activation, humoral immune responses, host defense, and immunological tolerance. We will also discuss the functional relationship between IgM-complement and IgM-FcμR pathways in regulating immunity and tolerance. Finally, we will discuss the potential involvement of FcμR in human diseases.

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Plants extensively employ leucine-rich repeat receptor-like kinases (LRR-RLKs), the largest family of RLKs, to control a wide range of growth and developmental processes as well as defense responses. To date, only a few direct downstream effectors for LRR-RLKs have been identified. We previously showed that the LRR-RLK EMS1 (EXCESS MICROSPOROCYTES1) and its ligand TPD1 (TAPETUM DETERMINANT1) are required for the differentiation of somatic tapetal cells and reproductive microsporocytes during early anther development in Here, we report the identification of β-carbonic anhydrases (βCAs) as the direct downstream targets of EMS1. EMS1 biochemically interacts with βCA proteins. Loss of function of genes caused defective tapetal cell differentiation, while overexpression of led to the formation of extra tapetal cells. EMS1 phosphorylates βCA1 at four sites, resulting in increased βCA1 activity. Furthermore, phosphorylation-blocking mutations impaired the function of βCA1 in tapetal cell differentiation; however, a phosphorylation mimic mutation promoted the formation of tapetal cells. βCAs are also involved in pH regulation in tapetal cells. Our findings highlight the role of βCA in controlling cell differentiation and provide insights into the posttranslational modification of carbonic anhydrases via receptor-like kinase-mediated phosphorylation.

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The marginal zone B cells (MZB) are located at the interface between the circulation and lymphoid tissue and as a gatekeeper play important roles in both innate and adaptive immune responses. We have previously found that MZB are significantly reduced in mice deficient in the IgM Fc receptor (FcμR) but how FcμR regulates the development and function of MZB remains unknown. In this study, we found that both marginal zone precursor (MZP) and MZB were decreased in FcμR−/− mice. The reduction of MZP and MZB was not due to impaired proliferation of these cells but rather due to their increased death. Further analysis revealed that FcμR−/− MZB had reduced tonic BCR signal, as evidenced by their decreased levels of phosphorylated SYK and AKT relative to WT MZB. MZB in FcμR−/− mice responded poorly to LPS in vivo when compared with MZB in WT mice. Consistent with the reduced proportion of MZB and their impaired response to LPS, antibody production against the type 1 T-independent Ag, NP-LPS, was significantly reduced in FcμR−/− mice. Moreover, FcμR−/− mice were highly susceptible to Citrobacter rodentium-induced sepsis. These results reveal a critical role for FcμR in the survival and activation of MZB and in protection against acute bacterial infection.