White blood cells called cytotoxic T cells (CTLs) play a crucial role in fighting disease; they identify other cells which are infected or damaged, then release toxic substances that destroy them. In recent years scientists have identified molecules within CTLs that help secrete the toxins. But little is known about the contrary process, which keeps levels of the substances low and prevents their release at the wrong time. A new study from the laboratories of Armin Rehm, Uta Höpken, and Bernd Dörken at the MDC/Charité University Hospital, along with several collaborating groups, has now exposed a new mechanism that helps cells control the toxins. One finding from the study, which appeared in the August 3 edition of the Journal of Clinical Investigation, is a potential link between the hormone estrogen and the way the immune system responds to cancer.
CTLs need to release enough toxins to cope with an infection, but secreting too many of them, at the wrong times, might damage healthy cells. They can even damage the cells that produce them, so the substances are carefully managed at each stage from synthesis to secretion. A part of this control involves packing the toxins into bubble-like compartments called vesicles. These membrane-wrapped packages are labeled with intricate protein "address labels" that dock onto partner molecules at the proper destination. They help fuse the vesicle to a larger compartment and deliver its contents.
Shortly after being made, CTL toxins are put into storage in compartments called secretory lysosomes. When time comes for secretion, the lysosomes are shuttled to the plasma membrane surrounding the cell. Each step provides an opportunity to control the process. The mechanism found by Rehm and his colleagues intervenes at an early stage, before toxins arrive at the secretory lysosomes.
The current findings follow work that the lab began several years ago on a protein called EBAG9. Many types of cells produce the molecule in modest amounts; it appears at much higher levels in some types of tumors, leading to its use as a diagnostic tool to determine whether some patients' tumors are likely to progress rapidly.. But the normal functions of EBAG9 in healthy cells – which might shed light on its role in cancer – were unknown. One way to find out would be to force healthy cells to produce too much of the molecule and see what happened.
In 2005 Constantin Rüder, a scientist in Rehm's lab, did this experiment and discovered that certain types of nerve cells were no longer releasing neurotransmitters. Like CTL toxins, these tiny molecules are packed into vesicles to be secreted from the plasma membrane. The study provided a hint about why the process might be disrupted: in neurons, EBAG9 binds to a protein called Snapin, which is involved in the routing and fusion of vesicles.
In the present study, Rüder took the opposite approach: he removed EBAG9 entirely from a strain of mouse. The animals' CTLs released higher amounts of toxins, hinting that the loss of the molecule was interfering with a control mechanism. White blood cells lacking EBAG9 built smaller secretory lysosomes. On the other hand, the compartments were being loaded with excessive amounts of one of the toxins. Rüder and his colleagues learned that this effect depends on interactions between EBAG9 and another protein called gamma2-adaptin. When EBAG9 is present, the two molecules work together to keep the amounts low.
What overall effect did the removal of EBAG9 have on the mouse immune system? The researchers discovered that the CTLs of the mice were better at destroying target cells than those with the protein. The animals also had a stronger "memory" of prior diseases – they mounted a quicker defense when infectious agents challenged the body a second time.
CTLs produce different amounts of EBAG9 at different times. This suggests, Rehm says, that the molecule forms part of a control system which gives the cells the ability to fine-tune the release of toxins. Previous studies have shown that stimulating cells with estrogen, for example, causes them to make more EBAG9 – which means that the hormone may act as a "volume control" for toxin secretion.
It will be important to work out the details of this mechanism, Rehm says, because many scientists believe that CTLs help protect the body against cancer. Their proper function may depend on the behavior of EBAG9, which seems abnormal in some tumors. One implication is that estrogen, which stimulates the protein's production, may yield an important tool for influencing the behavior of white blood cells in cancer and other diseases.
- Russ Hodge
Rüder C, Höpken UE, Wolf J, Mittrücker HW, Engels B, Erdmann B, Wollenzin S, Uckert W, Dörken B, Rehm A. The tumor-associated antigen EBAG9 negatively regulates the cytolytic capacity of mouse CD8+ T cells. J Clin Invest. 2009 Aug;119(8):2184-203. Epub 2009 Jul 20.