Public Thesis defense - LIBST

The plasma membrane aquaporin ZmPIP1;1 interacts with ZmPIP2s to increase the water membrane permeability of maize cells by Bruna Rayane TEODORO JUNQUEIRA

Pour l’obtention du grade de Docteur en sciences

Plasma membrane intrinsic proteins (PIPs) are aquaporins responsible for the facilitated diffusion of water and small solutes through plasma membrane (PM). PIP subfamily can be classified as PIP1 and PIP2. The monomer is the functional unity and they assemble as homo- or heterotetramers. PIP2s are efficient water channels and they localize in the PM, while most of PIP1s are retained in the endoplasmic reticulum (ER) and need to physically interact with PIP2s within heterotetramers to be relocalized to the PM, where they generally act as water channels.

ZmPIP1;1 is a ubiquitous and one of the highest expressed AQPs in maize. Previous studies showed that the co-expression of ZmPIP1;1 with ZmPIP2;5 in Xenopus oocytes did not lead to a synergistic effect on membrane water permeability. However, both isoforms physically interacted and were found in the PM. ZmPIP1;1 has been considered to be an inactive water channel until nowadays. In this thesis, we aimed at unraveling its role using different expression systems.

We firstly uncovered the role of two neighboring amino acid residues located in the loop E inhibiting the water channel activity when expressed in Xenopus oocytes. We generated an activated ZmPIP1;1 form and also introduced an additional mutation, W94A, known to block the water pore, thus confirming that ZmPIP1;1 could work as a water channel under specific loop E composition.

We also studied ZmPIP1;1 in maize Black Mexican Sweet (BMS) suspension cells. We overexpressed (OE) ZmPIP1;1 and the protein was detected in the PM. Surprisingly, the membrane water permeability of ZmPIP1;1 OE protoplasts was higher than the permeability of the wild-type (WT) protoplasts suggesting that either ZmPIP1;1 was transporting water by itself, or it was leading to an enrichment of active ZmPIP2s in the PM via heteromerization. The first hypothesis was tested by the generation of BMS lines expressing the inactive ZmPIP1;1W94A channel. This mutated cell lines exhibited a PM water permeability similar to the WT cells, suggesting that ZmPIP1;1 has an intrinsic ability to facilitate water transport in BMS cells. The second hypothesis was explored by PM purification of ZmPIP1;1 OE lines and quantification of ZmPIP2 proteins. Actually, a significant increase in the ZmPIP2 content in the PM was observed, possibly explaining the unexpected water permeability presented by these cell lines.

Altogether, our data uncovered the role of two amino acids in loop E responsible for the water channel inactivity of ZmPIP1;1. Also, BMS experiments suggested that ZmPIP1;1 may work as an active water channel in maize cells. Besides, ZmPIP1;1 was also able to regulate the water membrane permeability by increasing ZmPIP2 abundance in the PM via heteromerization. The mechanisms underlying this interaction, remains to be elucidated, for instance, with the structural determination of ZmPIP1 and ZmPIP2 heterotetramers.

Jury members :

• Prof. François Chaumont (UCLouvain), supervisor
• Prof. Michel Ghislain (UCLouvain), chairperson
• Prof. Pierre Morsomme (UCLouvain), secretary
• Prof. Charles Hachez (UCLouvain)
• Prof. Gerd Patrick Bienert (Technical University of Munich, Germany)
• Prof. Karina Alleva (University of Buenos Aires, Argentina)

Pay attention :

The public defense of Bruna Rayane Teodoro Junqueira scheduled for Friday 08 July at 02:00 p.m. will also take place in the form of a video conference

ID de réunion : 851 6774 7262

Code secret : 690142

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