Single Mutations in the Transmembrane Domains of Maize Plasma Membrane Aquaporins Affect the Activity of Monomers within a Heterotetramer

Aquaporins are channels facilitating the diffusion of water and/or small uncharged solutes ecross biological membranes. They assemble as homotetramers but some of them also form heterotetramers, especially in plants. In Zea mays, aquaporins belonging to the plasma membrane intrinsic protein （PIP） subfamily are clustered into two groups, PIP1 and PIP2, which exhibit different water-channel activities when expressed in Xenopus oocytes. When PIP1 and PIP2 isoforms are co-expressed, they physically interact to modulate their subcellular localization and channel activity. Here, we demonstrated by affinity chromatography purification that, when co-expressed in Xenopus oocytes, the maize PIP1;2 and PIP2;5 isoforms assemble as homo- and heterodimers within heterotetramers. We built the 3D structure of such heterotetramers by comparative modeling on the basis of the spinach SoPIP2;1 X-ray structure and identified amino acid residues in the transmembrane domains which putatively interact at the interfaces between monomers. Their roles in the water-channel activity, subcellular localization, protein abundance, and physical interac- tion were investigated by mutagenesis. We highlighted single-residue substitutions that either inactivated PIP2;5 or activated PIP1;2 without affecting their interaction. Interestingly, the Phe220Ala mutation in the transmembrane domain 5 of PIP1 ;2 activated its water-channel activity and, at the same time, inactivated PIP2;5 within a heterotetramer. Altogether, these data contribute to a better understanding of the interaction mechanisms between PIP isoforms and the role of heterotetramerization on their water-channel activity.

Annexin A2-S100A10 heterotetramer is upregulated by PML/RARα fusion protein and promotes plasminogen-dependent fibrinolysis and matrix invasion in acute promyelocytic leukemia

Aberrant expression of annexin A2-SI00A10 heterotetramer （AIIt） associated with PML/RARα fusion protein causes lethal hyperfibrinolysis in acute promyelocytic leukemia （APL）, but the mechanism is unclear. To facilitate the investigation of regulatory association between ANXA2 and promyelocytic leukemia/retinoic acid receptor α （PML/RARα） fusion protein, this work was performed to determine the transcription start site of ANXA2 promoter with rapid amplification of 5＇-cDNA ends analysis. Zinc-induced U937/PR9 cells expressed PML/RARα fusion protein, and resultant increases in ANXA2 transcripts and translational expressions of both ANXA2 and S100A10, while S100A10 transcripts remained constitutive. The transactivation of ANXA2 promoter by PML/RARα fusion protein was 3.29 ± 0.13 fold higher than that by control pSG5 vector or wild-type RARer. The overexpression of ANXA2 in U937 transfected with full-length ANXA2 cDNA was associated with increased S100A10 subunit, although S100A10 transcripts remained constitutive. The tPA-dependent initial rate of plasmin generation （IRPG） in zinc-treated U937/PR9 increased by 2.13-fold, and cell invasiveness increased by 27.6%. Antibodies against ANXA2, S100A10, or combination of both all remarkably inhibited the IRPG and invasiveness in U937/PR9 and NB4. Treatment of zinc-induced U937/PR9 or circulating APL blasts with all-trans retinoic acid （ATRA） significantly reduced cell surface ANXA2 and S100A10 and associated reductions in IRPG and invasiveness. Thus, PML/RARα fusion protein transactivated the ANXA2 promoter to upregulate ANXA2 and accumulate S100A10. Increased AIIt promoted IRPG and invasiveness, both of which were partly abolished by antibodies against ANXA2 and S100A10 or by ATRA.

Crystallographic Studies of Cephalosporin Acylase from Pseudomonas sp. Strain 130

The cephalosporin acylases are a group of enzymes that hydrolyze cephalosporin C and/or glutaryl 7 aminocephalosporanic acid to produce 7 aminocephalosporanic acid. The cephalosporin acylase from Pseudomonas sp. strain 130 was crystallized in two different forms suitable for structural studies. A tetragonal crystal form diffracted to 0.24 nm belonged to the space group P4 12 12. There was one αβ heterodimer per asymmetric unit. A second crystal form diffracted to 0.21 nm belonged to the space group P2 1. There was four αβ heterodimers per asymmetric unit. The tetragonal crystal structure of CA 130 was determined using the multiwavelength anomalous diffraction method and the P2 1 crystal structure was then determined using the molecular replacement method.

Inwardly rectifying potassium channel 5.1:Structure,function,and possible roles in diseases

Inwardly rectifying potassium(Kir)channels make it easier for K^(+)to enter into a cell and subsequently regulate cellular biological functions.Kir5.1(encoded by KCNJ16)alone can form a homotetramer and can form heterotetramers with Kir4.1(encoded by KCNJ10)or Kir4.2(encoded by KCNJ15).In most cases,homomeric Kir5.1 is non-functional,while hetero-meric Kir5.1 on the cell membrane contributes to the inward flow of K^(+)ions,which can be regulated by intracellular pH and a variety of signaling mechanisms.In the form of a heterotetramer,Kir5.1 regulates Kir4.1/4.2 activity and is involved in the maintenance of nephron function.Actually,homomeric Kir5.1 may also play a very important role in diseases,including in the ventilatory response to hypoxia and hypercapnia,hearing impairment,cardio-vascular disease and cancer.With an increase in the number of studies into the roles of Kir channels,researchers are paying more attention to the pathophysiological functions of Kir5.1.This minireview provides an overview regarding these Kir5.1 roles.