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Iodothyronine Interactions with the System L1 Amino Acid Exchanger in 3T3-L1 Adipocytes.

Mitchell FE, Roy LA, Taylor PM - J Thyroid Res (2010)

Bottom Line: Efflux of preloaded L-[(3)H]phenylalanine from 3T3-L1 adipocytes was trans stimulated by external leucine, demonstrating the obligatory exchange mechanism of System L1 transport.T(3) (10 μM) did not significantly trans stimulate L-[(3)H]phenylalanine efflux, but did competitively inhibit the trans stimulatory effect of 10 μM leucine.The results highlight strong competitive interactions between iodothyronines (T(3), rT(3)) and amino acids for transport by System L1 in adipocytes, which may impact cellular iodothyronine exchanges during altered states of protein nutrition.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Physiology, College of Life Sciences, James Black Centre, University of Dundee, Dundee DD1 5EH, UK.

ABSTRACT
Thyroid hormones enter isolated white adipocytes largely by a System L1-type amino acid transporter en route to exerting genomic actions. Differentiated 3T3-L1 mouse adipocytes in culture express mRNA for LAT1 (the catalytic subunit of high-affinity System L1). L-[(125)I]-T(3) uptake into 3T3-L1 adipocytes included a substantial saturable component inhibited by leucine. L-[(3)H]phenylalanine uptake into 3T3-L1 cells was saturable (K(m) of 31 μM), competitively inhibited by T(3) (K(i) of 1.2 μM) and blocked by leucine, BCH, and rT(3) as expected for substrate interactions of System L1. Efflux of preloaded L-[(3)H]phenylalanine from 3T3-L1 adipocytes was trans stimulated by external leucine, demonstrating the obligatory exchange mechanism of System L1 transport. T(3) (10 μM) did not significantly trans stimulate L-[(3)H]phenylalanine efflux, but did competitively inhibit the trans stimulatory effect of 10 μM leucine. The results highlight strong competitive interactions between iodothyronines (T(3), rT(3)) and amino acids for transport by System L1 in adipocytes, which may impact cellular iodothyronine exchanges during altered states of protein nutrition.

No MeSH data available.


Related in: MedlinePlus

Effect of amino acids and iodothyronines on uptake of phenylalanine into 3T3-L1 adipocytes. [3H]phenylalanine uptake was measured at 5 μM over 3 minutes in the absence (control) or presence of 10 μM iodothyronine or 10 mM amino acid in the uptake buffer. Values are expressed ± SEM of 3 measurements (except where indicated) as % of the control flux. **P < .01; value significantly different from control using Dunnett's Multiple Comparison Test.
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fig2: Effect of amino acids and iodothyronines on uptake of phenylalanine into 3T3-L1 adipocytes. [3H]phenylalanine uptake was measured at 5 μM over 3 minutes in the absence (control) or presence of 10 μM iodothyronine or 10 mM amino acid in the uptake buffer. Values are expressed ± SEM of 3 measurements (except where indicated) as % of the control flux. **P < .01; value significantly different from control using Dunnett's Multiple Comparison Test.

Mentions: The uptake of 5 μM phenylalanine into 3T3-L1 cells (0.465 ± 0.065 nmol·mg protein−1·min−1, n = 12) was saturable, being reduced more than 95% by addition of excess (10 mM) unlabelled phenylalanine, leucine or BCH (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid; a synthetic System L-type amino acid transport inhibitor) (Figure 2). 5 μM phenylalanine uptake was also substantially inhibited by T3 and rT3 (82% and 57% inhibition, resp.) at 10 μM, although inhibition by T4 (17%) and triiodothyroacetic acid (TRIAC; a T3 analogue lacking the α-amino grouping) did not achieve statistical significance at this concentration (Figure 2). Saturable phenylalanine uptake into 3T3-L1 cells had a Km of 31 μM and a Vmax of 2.6 ± 0.5 nmol·mg protein−1·min−1 (mean ± sem, n = 6; see Figure 3). The inhibitory effect of T3 on phenylalanine uptake also appeared to be competitive, with a Ki of the order of 1-2 μM (see Figure 4).


Iodothyronine Interactions with the System L1 Amino Acid Exchanger in 3T3-L1 Adipocytes.

Mitchell FE, Roy LA, Taylor PM - J Thyroid Res (2010)

Effect of amino acids and iodothyronines on uptake of phenylalanine into 3T3-L1 adipocytes. [3H]phenylalanine uptake was measured at 5 μM over 3 minutes in the absence (control) or presence of 10 μM iodothyronine or 10 mM amino acid in the uptake buffer. Values are expressed ± SEM of 3 measurements (except where indicated) as % of the control flux. **P < .01; value significantly different from control using Dunnett's Multiple Comparison Test.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2957699&req=5

fig2: Effect of amino acids and iodothyronines on uptake of phenylalanine into 3T3-L1 adipocytes. [3H]phenylalanine uptake was measured at 5 μM over 3 minutes in the absence (control) or presence of 10 μM iodothyronine or 10 mM amino acid in the uptake buffer. Values are expressed ± SEM of 3 measurements (except where indicated) as % of the control flux. **P < .01; value significantly different from control using Dunnett's Multiple Comparison Test.
Mentions: The uptake of 5 μM phenylalanine into 3T3-L1 cells (0.465 ± 0.065 nmol·mg protein−1·min−1, n = 12) was saturable, being reduced more than 95% by addition of excess (10 mM) unlabelled phenylalanine, leucine or BCH (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid; a synthetic System L-type amino acid transport inhibitor) (Figure 2). 5 μM phenylalanine uptake was also substantially inhibited by T3 and rT3 (82% and 57% inhibition, resp.) at 10 μM, although inhibition by T4 (17%) and triiodothyroacetic acid (TRIAC; a T3 analogue lacking the α-amino grouping) did not achieve statistical significance at this concentration (Figure 2). Saturable phenylalanine uptake into 3T3-L1 cells had a Km of 31 μM and a Vmax of 2.6 ± 0.5 nmol·mg protein−1·min−1 (mean ± sem, n = 6; see Figure 3). The inhibitory effect of T3 on phenylalanine uptake also appeared to be competitive, with a Ki of the order of 1-2 μM (see Figure 4).

Bottom Line: Efflux of preloaded L-[(3)H]phenylalanine from 3T3-L1 adipocytes was trans stimulated by external leucine, demonstrating the obligatory exchange mechanism of System L1 transport.T(3) (10 μM) did not significantly trans stimulate L-[(3)H]phenylalanine efflux, but did competitively inhibit the trans stimulatory effect of 10 μM leucine.The results highlight strong competitive interactions between iodothyronines (T(3), rT(3)) and amino acids for transport by System L1 in adipocytes, which may impact cellular iodothyronine exchanges during altered states of protein nutrition.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Physiology, College of Life Sciences, James Black Centre, University of Dundee, Dundee DD1 5EH, UK.

ABSTRACT
Thyroid hormones enter isolated white adipocytes largely by a System L1-type amino acid transporter en route to exerting genomic actions. Differentiated 3T3-L1 mouse adipocytes in culture express mRNA for LAT1 (the catalytic subunit of high-affinity System L1). L-[(125)I]-T(3) uptake into 3T3-L1 adipocytes included a substantial saturable component inhibited by leucine. L-[(3)H]phenylalanine uptake into 3T3-L1 cells was saturable (K(m) of 31 μM), competitively inhibited by T(3) (K(i) of 1.2 μM) and blocked by leucine, BCH, and rT(3) as expected for substrate interactions of System L1. Efflux of preloaded L-[(3)H]phenylalanine from 3T3-L1 adipocytes was trans stimulated by external leucine, demonstrating the obligatory exchange mechanism of System L1 transport. T(3) (10 μM) did not significantly trans stimulate L-[(3)H]phenylalanine efflux, but did competitively inhibit the trans stimulatory effect of 10 μM leucine. The results highlight strong competitive interactions between iodothyronines (T(3), rT(3)) and amino acids for transport by System L1 in adipocytes, which may impact cellular iodothyronine exchanges during altered states of protein nutrition.

No MeSH data available.


Related in: MedlinePlus