Possible functions

To date there is no published data for MMD and MMD2 protein functions. In mouse BMM, the rapid increase of mMMD transcript expression upon LPS stimulation suggests an association of the protein with the first phase of the immune response. This latter involves pro-inflammatory processes and the activation of transcription factors such as NFκB, which in turn promotes gene transcription and expression. It is not known whether the increase in mMMD expression is directly induced by NFκB or by other unknown factors, or whether it

contributes to the inflammatory response mediated by NFκB (Abul K.Abbas & Andrew H.Lichtman, 2003).

A second hint involving MMD in activated macrophages processes was provided by Sawcer S. et al. (Sawcer et al., 2002), who classified hMMD as a susceptibility gene for Multiple Sclerosis (MS). MS is a chronic inflammatory demyelinating disease of the central nervous system, resulting in impaired nerve conduction. The keys to the pathophysiology are activated lymphocytes, macrophages, astrocytes and microglia. Activated microglia/macrophages are involved in the demyelination and secrete inflammatory cytokines such as TNFα, IL-1 and IL-6 which additionally contribute to the formation of free radicals and tissue damage (Burke B & Lewis Claire E., 2002). The involvement of hMMD in these processes suggests either a role in inflammation or as protector against brain damage.

Recently, Lünemann et al. (Lunemann et al., 2006) showed that MAF/MMD (rat MMD orthologue) was upregulated in regions of the brain after an entorhinal cortex lesion (ECL).

ECL models post-traumatic tissue remodeling of neuronal structure and connectivity (Lunemann et al., 2006). The early phase of this process is characterized by the recruitment, into the lesion site, of inflammatory cells such as microglia and newly infiltrated macrophages, whichs produce IL-1β. In the late phase of ECL (two days later), tissue repair and remodeling are initiated, where microglia were shown to play a crucial role (Mabuchi et al., 2000). MMD/MAF expression in this latter stage of ECL suggests its role in tissue remodeling rather than inflammation (Lunemann et al., 2006). However, Lünemann et al. did not investigate whether MAF/MMD was also expressed during the early phase of ECL.

Taken together, MMD function may be associated with immediate post-stimulatory activation of macrophages, and is likely associated with tissue remodeling.

The recent classification of MMD and MMD2 into PAQRs family (Tang et al., 2005) shed more light into their putative functions, however few clues are available linking MMD proteins to these receptors.

On the transcript level, Northern blot analysis showed that both MMD and AdipoRs mRNA were expressed in macrophages but also in liver, skeletal muscle, adipose tissue and placenta (Yamauchi et al., 2003a). On the other hand, MMD2 shared with mPRα (PAQR7) the expression in testis and with mPRβ and MMD the expression in brain. However, apart from

the similar expression pattern, there is no other evidence which relates functionally MMD proteins to PAQRs.

Interestingly, in the placenta in addition to hMMD and AdipoRs (Yamauchi et al., 2003a), mPRα and PAQR9 were expressed (Fernandes et al., 2005; Zhu et al., 2003a) and functionally associated with the onset of pregnancy. PAQR3 was also strongly and constantly expressed in the endometrium during the cycle and upon pregnancy, where it was predicted to responsible for the maintenance of the tissue homeostasis (Fernandes et al., 2005). In fact, in the early phases of pregnancy, a balance between pro- and anti-oxidant factors is important in the placenta microenvironment for the trophoblast invasion (Biondi et al., 2005). Macrophages, which are in close contact with trophoblasts, play a pivotal role in this process by supporting a certain grade of inflammation (Mor & Abrahams, 2003; Zhu et al., 1997). Interestingly, PAQR3 orthologue in Drosophila melanogaster, CG7530, was identified as a resistant gene against oxidative stress (Monnier et al., 2002). It is therefore likely that PAQR3 may play the same role in the placenta during the pregnancy. The next question would be if hMMD can also be involved in the onset of pregnancy and in oxidative stress as well.

On the protein level, PAQRs differ in their subcellular localization. PAQR1 and PAQR2 (Yamauchi et al., 2003a) as well as the yeast YOL002c (Narasimhan et al., 2005) were defined as plasma membrane proteins (Zhu et al., 2003b). PAQR8 was located in lysosomes (Suzuki et al., 2001) and only mPRα (PAQR7) shares with mMMD the perinuclear and nuclear membrane localization (Fernandes et al., 2005). However, amino acid residues with a predicted metal-binding property were found conserved in all PAQRs (see section 6.2) (Tang et al., 2005; Lyons et al., 2004).

In the yeast PAQR orthologue, YOL002c, the metal-binding amino acid residues were associated with zinc metabolism. YOL002c, also termed “Implicated in Zinc Homeostasis”

(IZH)-2 was found to be induced by a zinc limited environment. Its function was also associated with lipid metabolism (Karpichev et al., 2002). Additionally, it has been speculated that IZH-2 could be an ion channel (Lyons et al., 2004).

In AdipoRs and mMMD proteins, where the membrane topology was determined, the metal-binding residues, located outside the TM regions, face the cytosol. It is therefore tempting to propose that both proteins may be implicated in the cellular zinc homeostasis.

An ion channel property was proposed for the human PAQR8 (mPRβ, also termed as lysosomal membrane protein brain (LMPB)-1). PAQR8 was associated with a brain

pathology, namely juvenile myoclonic epilepsy (JME), which is characterized by seizures without detectable brain lesions (Suzuki et al., 2001). Defects in ion channel proteins were implicated in JME.

One can assume that MMD proteins could be an ion channels as well. The fact that they were defined as related to Hly-III (Rehli et al., 1995; Tang et al., 2005), which was shown to form membrane pores in erythrocytes (Baida & Kuzmin, 1996), may support this assumption.

Taken together, MMD and MMD2 expressed in different tissues, are likely to be involved in different processes. Mouse MMD intracellular localization and topology suggest a putative role in maintenance of the cellular homeostasis. Whether the metal-binding property of the conserved amino acid residues plays a role in oxidative stress, or whether MMD protein are ion channels, remains to be determined.