Characterization of the small Arabidopsis thaliana GTPase and ADP-ribosylation factor-like 2 protein TITAN5
Small GTPases comprise key proteins in signal transduction that function by conformational switching ability between GDP- and GTP-bound states. The ADP-ribosylation factor (ARF) family is involved in vesicle trafficking and cellular functions. Though evolutionarily well conserved, little is known about ARF and ARF-like GTPases in plants. Here, we characterized functional properties and cellular localization of the essential small ARF-like GTPase TITAN5/HALLIMASCH/ARL2/ARLC1 (hereafter termed TTN5) from Arabidopsis thaliana. TTN5 showed rapid guanine nucleotide exchange capacity comparable to that of human counterparts, but a remarkably low GTP hydrolysis reaction. A TTN5Q70L mutant had enhanced nucleotide exchange activity, indicative of intracellular activation, while TTN5T30N with fast nucleotide dissociation can be considered a dominant-negative form. This suggests that TTN5 is present in GTP-loaded active form in the cells. YFP-tagged TTN5 and the two derived mutant variants were located at multiple sites of the endomembrane system in the epidermis of Arabidopsis seedlings and Nicotiana benthamiana leaves. While YFP-TTN5 and YFP-TTN5Q70L were highly mobile in the cells, mobility was reduced for TTN5T30N. Colocalization with endomembrane markers in combination with pharmacological treatments resolved localization at membrane sites and showed that YFP-TTN5 and YFP-TTN5Q70L were located in Golgi stacks, multivesicular bodies, while this was less the case for YFP-TTN5T30N. On the other hand, all three TTN5 forms were located at the plasma membrane. Hence, the unusual capacity of rapid nucleotide exchange activity of the small ARF-like GTPase TTN5 is linked with cell membrane dynamics, likely associated with vesicle transport pathways in the endomembrane system.
| Medienart: |
E-Artikel |
|---|
| Erscheinungsjahr: |
2023 |
|---|---|
| Erschienen: |
2023 |
| Enthalten in: |
Zur Gesamtaufnahme - year:2023 |
|---|---|
| Enthalten in: |
bioRxiv : the preprint server for biology - (2023) vom: 05. Mai |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Mohr, Inga [VerfasserIn] |
|---|
| Links: |
|---|
| Themen: |
ARF-like |
|---|
| Anmerkungen: |
Date Revised 19.10.2023 published: Electronic Citation Status PubMed-not-MEDLINE |
|---|
| doi: |
10.1101/2023.04.27.538563 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM356657957 |
|---|
| LEADER | 01000naa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM356657957 | ||
| 003 | DE-627 | ||
| 005 | 20231226070928.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231226s2023 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.1101/2023.04.27.538563 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n1188.xml |
| 035 | |a (DE-627)NLM356657957 | ||
| 035 | |a (NLM)37162876 | ||
| 035 | |a (PII)2023.04.27.538563 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Mohr, Inga |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Characterization of the small Arabidopsis thaliana GTPase and ADP-ribosylation factor-like 2 protein TITAN5 |
| 264 | 1 | |c 2023 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Revised 19.10.2023 | ||
| 500 | |a published: Electronic | ||
| 500 | |a Citation Status PubMed-not-MEDLINE | ||
| 520 | |a Small GTPases comprise key proteins in signal transduction that function by conformational switching ability between GDP- and GTP-bound states. The ADP-ribosylation factor (ARF) family is involved in vesicle trafficking and cellular functions. Though evolutionarily well conserved, little is known about ARF and ARF-like GTPases in plants. Here, we characterized functional properties and cellular localization of the essential small ARF-like GTPase TITAN5/HALLIMASCH/ARL2/ARLC1 (hereafter termed TTN5) from Arabidopsis thaliana. TTN5 showed rapid guanine nucleotide exchange capacity comparable to that of human counterparts, but a remarkably low GTP hydrolysis reaction. A TTN5Q70L mutant had enhanced nucleotide exchange activity, indicative of intracellular activation, while TTN5T30N with fast nucleotide dissociation can be considered a dominant-negative form. This suggests that TTN5 is present in GTP-loaded active form in the cells. YFP-tagged TTN5 and the two derived mutant variants were located at multiple sites of the endomembrane system in the epidermis of Arabidopsis seedlings and Nicotiana benthamiana leaves. While YFP-TTN5 and YFP-TTN5Q70L were highly mobile in the cells, mobility was reduced for TTN5T30N. Colocalization with endomembrane markers in combination with pharmacological treatments resolved localization at membrane sites and showed that YFP-TTN5 and YFP-TTN5Q70L were located in Golgi stacks, multivesicular bodies, while this was less the case for YFP-TTN5T30N. On the other hand, all three TTN5 forms were located at the plasma membrane. Hence, the unusual capacity of rapid nucleotide exchange activity of the small ARF-like GTPase TTN5 is linked with cell membrane dynamics, likely associated with vesicle transport pathways in the endomembrane system | ||
| 650 | 4 | |a Preprint | |
| 650 | 4 | |a ARF-like | |
| 650 | 4 | |a ARL2 | |
| 650 | 4 | |a GTPase | |
| 650 | 4 | |a TTN5 | |
| 650 | 4 | |a endomembrane | |
| 650 | 4 | |a plasma membrane | |
| 650 | 4 | |a vesicle | |
| 700 | 1 | |a Mirzaiebadizi, Amin |e verfasserin |4 aut | |
| 700 | 1 | |a Sanyal, Sibaji K |e verfasserin |4 aut | |
| 700 | 1 | |a Chuenban, Pichaporn |e verfasserin |4 aut | |
| 700 | 1 | |a Ahmadian, Mohammad R |e verfasserin |4 aut | |
| 700 | 1 | |a Ivanov, Rumen |e verfasserin |4 aut | |
| 700 | 1 | |a Bauer, Petra |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t bioRxiv : the preprint server for biology |d 2020 |g (2023) vom: 05. Mai |w (DE-627)NLM31090014X |7 nnns |
| 773 | 1 | 8 | |g year:2023 |g day:05 |g month:05 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1101/2023.04.27.538563 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |j 2023 |b 05 |c 05 | ||